PhD Thesis Notes: Insights on ATRA-Sensitivity Cluster

INSIGHTS ON PHD THESIS RESULTS 8.1 Cluster of 42 Genes Correlated with ATRA-Sensi vity: Posi ve Correla on: 1. EGF (Epidermal Growth Factor): o Func on: EGF is a growth factor that s mulates cell growth, prolifera on, and diﬀeren a on by binding to its receptor, EGFR. It plays a cri cal role in wound healing and is implicated in the progression of various cancers due to its ability to promote cell division and survival. 2. HBE1 (Hemoglobin Subunit Epsilon 1): o Func on: HBE1 is part of the embryonic hemoglobin complex, primarily expressed during the early stages of fetal development. It is eventually replaced by adult hemoglobin components, such as alpha and beta globin, as development progresses. 3. OR52D1 (Olfactory Receptor Family 52 Subfamily D Member 1): o Func on: This gene encodes an olfactory receptor involved in the detec on of odorants. Olfactory receptors are part of a large family of G-protein-coupled receptors (GPCRs) and play a crucial role in the sense of smell by detec ng chemical s muli in the environment. 4. OR51B5 (Olfactory Receptor Family 51 Subfamily B Member 5): o Func on: OR51B5 encodes another olfactory receptor, contribu ng to the diversity of odor detec on in humans. It is involved in the signal transduc on pathways that convert chemical signals (odors) into neural signals. 5. KCNJ8 (Potassium Inwardly Rec fying Channel Subfamily J Member 8): o Func on: KCNJ8 encodes a component of ATP-sensi ve potassium channels (KATP channels), which regulate membrane poten al and play a crucial role in various physiological processes, including cardiac rhythm, vascular tone, and insulin release. 6. ERBB4 (Erb-B2 Receptor Tyrosine Kinase 4): o Func on: ERBB4 is a member of the epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases. It is involved in cellular processes such as diﬀeren a on, prolifera on, and apoptosis. ERBB4 signaling is essen al in the development of the heart, nervous system, and mammary glands. 7. WNT2 (Wnt Family Member 2): o Func on: WNT2 encodes a protein involved in the Wnt signaling pathway, which regulates crucial aspects of cell fate determina on, cell migra on, cell polarity, and organogenesis during embryonic development. It is also implicated in cancer development. 8. OR51I2 (Olfactory Receptor Family 51 Subfamily I Member 2): o Func on: This gene encodes an olfactory receptor, contribu ng to the ability to detect and diﬀeren ate a wide range of odors. These receptors are involved in the ini al steps of the olfactory signal transduc on pathway. 9. SPX (Spexin Hormone): o Func on: Spexin is a pep de hormone involved in regula ng appe te, energy metabolism, and fat storage. It has been studied for its role in obesity and metabolic disorders and is also involved in cardiovascular func ons. 10. FLNC (Filamin C): Func on: Filamin C is an ac n-binding protein that plays a cri cal role in maintaining the structural integrity of the cytoskeleton, par cularly in muscle cells. It is involved in linking cell membrane components to the ac n cytoskeleton and is crucial for muscle cell stability and func on. 11. COL6A1 (Collagen Type VI Alpha 1 Chain): Func on: COL6A1 encodes a component of type VI collagen, a part of the extracellular matrix that provides structural support to ssues. It plays a role in the stability and organiza on of collagen ﬁbrils and is important in maintaining the integrity of muscle, skin, and other connec ve ssues. Pag. 147 a 170 12. OR51J1 (Olfactory Receptor Family 51 Subfamily J Member 1): Func on: This gene encodes an olfactory receptor involved in odor detec on. Like other olfactory receptors, it contributes to the sensory percep on of smell by detec ng speciﬁc odor molecules and triggering a signal transduc on cascade. 13. OR51I1 (Olfactory Receptor Family 51 Subfamily I Member 1): Func on: OR51I1 encodes an olfactory receptor that par cipates in the detec on of odors. These receptors are key players in the olfactory system, allowing organisms to detect and respond to environmental chemical cues. 14. PITX2 (Paired Like Homeodomain 2): Func on: PITX2 is a transcrip on factor involved in the development of the le -right asymmetry in the body, as well as the development of various organs, including the eyes, heart, and teeth. Muta ons in PITX2 can lead to developmental disorders such as Rieger syndrome. 15. SIRT3 (Sirtuin 3): Func on: SIRT3 is a member of the sirtuin family of NAD+-dependent deacetylases, primarily located in the mitochondria. It plays a cri cal role in regula ng mitochondrial func on, energy metabolism, and oxida ve stress, and is involved in aging and metabolic regula on. 16. RTP2 (Receptor Transporter Protein 2): Func on: RTP2 facilitates the transport of olfactory receptors to the cell membrane, where they can func on in odor detec on. It is important for the proper func oning of the olfactory system by ensuring that receptors are correctly localized. 17. EHD2 (EH Domain Containing 2): Func on: EHD2 is involved in endocy c recycling and membrane traﬃcking. It plays a role in the regula on of plasma membrane dynamics, including the forma on and traﬃcking of caveolae, which are small invagina ons in the cell membrane. 18. HBG1 (Hemoglobin Subunit Gamma 1): Func on: HBG1 encodes a gamma globin subunit that is part of fetal hemoglobin (HbF). HbF has a higher aﬃnity for oxygen than adult hemoglobin and is cri cal for oxygen transport during fetal development. It is replaced by adult hemoglobin a er birth. 19. TLL1 (Tolloid Like 1): Func on: TLL1 encodes a metalloprotease involved in the processing of extracellular matrix proteins and is essen al for heart development. It plays a role in ssue remodeling during embryogenesis and is cri cal for the proper forma on of cardiac structures. 20. LOXL1 (Lysyl Oxidase Like 1): Func on: LOXL1 encodes an enzyme involved in the crosslinking of collagen and elas n ﬁbers in the extracellular matrix, contribu ng to the stability and elas city of connec ve ssues. It is associated with the development of diseases like exfolia on syndrome, which aﬀects the eyes. 21. PAX9 (Paired Box 9): Func on: PAX9 is a transcrip on factor that plays a key role in the development of teeth, vertebrae, and other skeletal structures during embryogenesis. Muta ons in PAX9 are associated with tooth agenesis, where some teeth fail to develop. 22. NUP98 (Nucleoporin 98): Func on: NUP98 encodes a component of the nuclear pore complex, which is essen al for the transport of molecules between the nucleus and the cytoplasm. NUP98 is involved in regula ng the ﬂow of gene c informa on and is implicated in certain types of leukemia due to its involvement in chromosomal transloca ons. 23. TPM1 (Tropomyosin 1): Func on: TPM1 encodes an ac n-binding protein that is integral to the regula on of muscle contrac on. It stabilizes ac n ﬁlaments and interacts with other proteins, such as troponin, to control the accessibility of myosin to the ac n ﬁlaments, enabling contrac on in muscle cells. Pag. 148 a 170 24. MEOX1 (Mesenchyme Homeobox 1): Func on: MEOX1 is a transcrip on factor that plays a role in the development of the paraxial mesoderm, which gives rise to skeletal muscles, vertebrae, and other structures. It is cri cal for the proper forma on of the somites during embryogenesis. 25. CPS1 (Carbamoyl-Phosphate Synthetase 1): Func on: CPS1 is an enzyme involved in the urea cycle, a cri cal metabolic pathway that detoxiﬁes ammonia by conver ng it to urea in the liver. Muta ons in CPS1 can lead to hyperammonemia, a condi on characterized by elevated levels of ammonia in the blood. 26. OR51B4 (Olfactory Receptor Family 51 Subfamily B Member 4): Func on: OR51B4 is another gene encoding an olfactory receptor, part of the large family of GPCRs involved in detec ng odor molecules. These receptors are responsible for ini a ng the sense of smell by interac ng with odorants and transmi ng signals to the brain. Nega ve Correla on: 27. ALX3 (ALX Homeobox 3): Func on: ALX3 encodes a transcrip on factor involved in craniofacial development. It is part of the homeobox gene family, which plays a crucial role in the regula on of genes during embryonic development, par cularly in the forma on of facial structures. 28. CHRM2 (Cholinergic Receptor Muscarinic 2): Func on: CHRM2 encodes a G-protein-coupled receptor (GPCR) that mediates the eﬀects of acetylcholine in the parasympathe c nervous system. It is involved in various physiological processes, including heart rate regula on, smooth muscle contrac on, and cogni ve func ons. 29. GRK2 (G Protein-Coupled Receptor Kinase 2): Func on: GRK2 is a kinase that phosphorylates ac vated GPCRs, leading to their desensi za on and internaliza on. This regula on is cri cal for controlling GPCR signaling, which is involved in numerous physiological processes such as neurotransmission, hormone response, and immune func on. 30. OR1N2 (Olfactory Receptor Family 1 Subfamily N Member 2): Func on: This gene encodes an olfactory receptor involved in the detec on of speciﬁc odor molecules. OR1N2 contributes to the wide range of olfactory receptors that allow the detec on of a diverse array of odors. 31. SRRM1 (Serine/Arginine Repe ve Matrix 1): Func on: SRRM1 is involved in the splicing of pre-mRNA, a crucial step in gene expression where introns are removed, and exons are joined together. SRRM1 plays a role in the regula on of alterna ve splicing, aﬀec ng the diversity of proteins produced by genes. 32. CLTB (Clathrin Light Chain B): Func on: CLTB encodes a component of clathrin, a protein that plays a cri cal role in the forma on of clathrin-coated vesicles. These vesicles are involved in processes such as endocytosis, where they mediate the internaliza on of receptors and other molecules from the cell surface. 33. BHMT (Betaine-Homocysteine S-Methyltransferase): Func on: BHMT is an enzyme involved in the metabolism of homocysteine, conver ng it to methionine using betaine as a methyl donor. This process is important in methyla on reac ons, which are crucial for DNA synthesis, repair, and gene regula on. 34. SIX6 (SIX Homeobox 6): Func on: SIX6 encodes a homeobox transcrip on factor involved in the development of the eye and the pituitary gland. It plays a crucial role in the regula on of genes during embryonic development, par cularly in the forma on of the re na and anterior pituitary. 35. UPF3A (UPF3 Regulator of Nonsense Mediated mRNA Decay A): Func on: UPF3A is involved in the nonsense-mediated mRNA decay (NMD) pathway, a surveillance mechanism that degrades mRNAs containing premature stop codons. This process prevents the produc on of truncated, poten ally harmful proteins. Pag. 149 a 170 36. ING1 (Inhibitor of Growth Family Member 1): Func on: ING1 encodes a tumor suppressor protein that plays a role in regula ng cell cycle progression, apoptosis, and chroma n remodeling. It is involved in the cellular response to DNA damage and has been implicated in the suppression of tumor growth. 37. OR4F21 (Olfactory Receptor Family 4 Subfamily F Member 21): Func on: OR4F21 is part of the olfactory receptor family, contribu ng to the detec on of odorants. Like other olfactory receptors, it is involved in the sense of smell by binding to speciﬁc molecules and ini a ng a signaling cascade that results in odor percep on. 38. FIP1L1 (Factor Interac ng with PAPOLA and CPSF1 Like 1): Func on: FIP1L1 is involved in the regula on of polyadenyla on, a process that adds a poly(A) tail to the 3' end of mRNA molecules, inﬂuencing their stability, transport, and transla on. FIP1L1 is also implicated in a speciﬁc fusion gene associated with hypereosinophilic syndrome, a disorder characterized by the overproduc on of eosinophils. 39. TLE3 (Transducin-Like Enhancer of Split 3): Func on: TLE3 encodes a transcrip onal co-repressor that interacts with various transcrip on factors to regulate gene expression during development. It is involved in processes such as neuronal development, adipogenesis, and the regula on of immune responses. 40. OR52M1 (Olfactory Receptor Family 52 Subfamily M Member 1): Func on: OR52M1 encodes an olfactory receptor that detects chemical s muli involved in the sense of smell. It belongs to the GPCR family and plays a role in ini a ng olfactory signal transduc on upon binding to speciﬁc odorants. 41. HBM (Hemoglobin Subunit Mu): Func on: HBM encodes a minor hemoglobin subunit expressed during early stages of human development. It is part of the embryonic and fetal hemoglobin complexes, which have a higher aﬃnity for oxygen compared to adult hemoglobin. 42. NRAP (Nebulin Related Anchoring Protein): Func on: NRAP is involved in the organiza on of ac n ﬁlaments in muscle cells, playing a cri cal role in the structural integrity and func on of sarcomeres, the basic units of muscle contrac on. It is essen al for the proper assembly and maintenance of the myoﬁbrillar structure in cardiac and skeletal muscles. Pag. 150 a 170 8.2 STRING (Search Tool for the Retrieval of Interac ng Genes/Proteins) Interac ons: 1. Predicted Interac ons: Gene Neighborhood (Green Line): o Deﬁni on: This interac on is based on the observa on that genes that are located near each other on the chromosome (in the same neighborhood) are o en func onally related. This proximity suggests that they may be involved in the same pathway or biological process. o Use in STRING: This type of interac on is par cularly relevant for prokaryo c organisms, where genes involved in the same pathway are o en organized in operons. Gene Fusions (Red Line): o Deﬁni on: Gene fusions occur when two or more genes that were originally separate in an ancestor species are found as a single, fused gene in another species. This fusion suggests that the products of these genes interact or func on together. o Use in STRING: Detec ng gene fusions can provide strong evidence that the proteins encoded by these genes interact or par cipate in a common func on. Gene Co-Occurrence (Blue Line): o Deﬁni on: This interac on is based on the observa on that certain genes are consistently found together across diﬀerent species. If two genes are o en present or absent together in diﬀerent genomes, it suggests that they might be func onally linked or part of the same biological process. o Use in STRING: Co-occurrence is o en used as an indicator of func onal associa on between proteins. 2. Known Interac ons: From Curated Databases (Light Blue Line): o Deﬁni on: These interac ons are derived from manually curated databases where interac ons have been conﬁrmed and documented based on exis ng literature. This type of interac on has high reliability because it is based on previously validated experiments. o Use in STRING: STRING integrates known interac ons from various curated databases to provide a comprehensive interac on network based on experimental evidence. Experimentally Determined (Pink Line): o Deﬁni on: These interac ons are iden ﬁed through experimental methods, such as yeast two- hybrid screening, co-immunoprecipita on, and other biochemical assays. The interac ons have been conﬁrmed in a lab se ng, making them highly reliable. o Use in STRING: Experimentally determined interac ons form a cri cal part of the network and are used as a founda on for predic ng further interac ons. 3. Others: Text Mining (Yellow-Green Line): o Deﬁni on: This method involves scanning scien ﬁc literature for co-occurrences of gene or protein names within the same sentence or abstract. If two genes or proteins are frequently men oned together in the literature, this suggests a poten al interac on. o Use in STRING: While text mining can suggest possible interac ons, these are typically treated with cau on because co-occurrence in text does not necessarily imply a direct interac on. Co-Expression (Black Line): o Deﬁni on: This interac on is based on the observa on that genes or proteins that are co-expressed (i.e., their expression levels rise and fall together across diﬀerent condi ons, ssues, or stages) are o en func onally related or part of the same pathway. o Use in STRING: Co-expression data is used to infer func onal associa ons, par cularly in cases where direct physical interac ons might not be evident. Protein Homology (Purple Line): o Deﬁni on: This refers to interac ons predicted based on the similarity between proteins (homologs) in diﬀerent species. If two proteins are homologous, they might perform similar func ons or interact in similar ways across species. o Use in STRING: Homology is used as an addi onal layer of evidence, par cularly when direct interac on data is not available for a par cular protein. Pag. 151 a 170 8.3 Tissue Development Pathways (Cluster PITX2, PAX9, ALX3, MEOX1, SIX6, TLE3): Tissue development pathways refer to the complex series of molecular and cellular processes that guide the forma on, growth, diﬀeren a on, and matura on of ssues in mul cellular organisms. These pathways involve the coordinated expression of genes, signaling molecules, and regulatory networks that ensure ssues develop with the correct structure and func on during embryogenesis, growth, and repair processes. The genes PITX2, PAX9, ALX3, MEOX1, SIX6, and TLE3 play crucial roles in various ssue development pathways. These genes encode transcrip on factors or other regulatory proteins that guide the development and diﬀeren a on of speciﬁc ssues during embryogenesis and later in life. Here’s an overview of each gene and its involvement in ssue development: 1. PITX2 (Paired-like homeodomain transcrip on factor 2): Role in Development: PITX2 is a transcrip on factor cri cal for the development of various ssues, including the eyes, teeth, heart, and lateral body asymmetry. Key Pathways: o Le -Right Asymmetry: PITX2 is a key gene in establishing le -right asymmetry during embryonic development. It is asymmetrically expressed on the le side of the developing embryo and is involved in the proper forma on of the heart and other organs. o Ocular Development: PITX2 plays a role in eye development, par cularly in the forma on of the anterior segment of the eye. Muta ons in PITX2 are associated with Axenfeld-Rieger syndrome, which aﬀects the development of the eyes, teeth, and umbilicus. 2. PAX9 (Paired box gene 9): Role in Development: PAX9 is a transcrip on factor involved in the development of the teeth, palate, and other craniofacial structures. Key Pathways: o Tooth Development: PAX9 is essen al for the development of molars and other teeth. It regulates the expression of genes involved in odontogenesis (tooth forma on) and is cri cal for the forma on of tooth buds and subsequent dental structures. o Craniofacial Development: PAX9 is also involved in the development of the palate and other craniofacial structures. Muta ons in PAX9 can lead to dental agenesis (missing teeth) and other craniofacial abnormali es. 3. ALX3 (ALX homeobox 3): Role in Development: ALX3 is a homeobox transcrip on factor involved in craniofacial and limb development. Key Pathways: o Craniofacial Development: ALX3 plays a role in the forma on of facial bones and ssues. It is part of a family of ALX genes that are important for craniofacial paterning and development. o Limb Development: ALX3 is also involved in limb development, par cularly in the regula on of paterning and outgrowth during embryogenesis. Muta ons in ALX3 can result in craniofacial dysmorphisms and limb anomalies. 4. MEOX1 (Mesenchyme homeobox 1): Role in Development: MEOX1 is a homeobox gene that plays a cri cal role in somitogenesis, the process that gives rise to the segmented structure of the vertebrate body. Key Pathways: o Somite Forma on: MEOX1 is crucial for the forma on and diﬀeren a on of somites, which are blocks of mesoderm that develop into the vertebrae, skeletal muscles, and dermis. It regulates the expression of genes involved in the development of these ssues. o Skeletal Muscle Development: MEOX1 is involved in the diﬀeren a on of myogenic progenitor cells into skeletal muscle cells. It plays a role in the early stages of muscle forma on during embryogenesis. Pag. 152 a 170 5. SIX6 (SIX homeobox 6): Role in Development: SIX6 is a homeobox gene involved in the development of the eye and brain. Key Pathways: o Eye Development: SIX6 is crucial for the development of the re na and other structures in the eye. It is part of the SIX family of genes, which are involved in the regula on of eye forma on and development. o Brain Development: SIX6 also plays a role in the development of the hypothalamus and other regions of the brain. It is involved in the regula on of neurogenesis and the forma on of speciﬁc neuronal popula ons. 6. TLE3 (Transducin-like enhancer of split 3): Role in Development: TLE3 is a transcrip onal corepressor involved in various developmental processes, including the regula on of diﬀeren a on and cell fate decisions. Key Pathways: o Wnt Signaling Pathway: TLE3 acts as a corepressor in the Wnt signaling pathway, which is crucial for the regula on of cell prolifera on, diﬀeren a on, and ssue paterning during development. o Adipogenesis: TLE3 has been implicated in the regula on of adipocyte diﬀeren a on, playing a role in the forma on of fat ssues during development and in adult ssue homeostasis. 8.4 WNT Pathway (Cluster WNT2, TLE3, EGF, ERBB3): The WNT signaling pathway is a complex network of proteins known for its roles in regula ng cell-to-cell interac ons during embryogenesis, ssue development, and homeostasis in adult ssues. The pathway is named a er the WNT gene, which encodes a family of secreted signaling proteins. WNT signaling is crucial for processes such as cell prolifera on, diﬀeren a on, migra on, and apoptosis. Dysregula on of the WNT pathway is associated with various diseases, including cancer, neurodegenera ve disorders, and congenital anomalies. There are three main branches of WNT signaling: 1. Canonical (β-catenin dependent) WNT Pathway: This pathway involves the stabiliza on of β-catenin, which then translocates to the nucleus to regulate the transcrip on of target genes. 2. Non-Canonical WNT Pathway (Planar Cell Polarity Pathway): This pathway regulates the cytoskeleton and aﬀects cell movement and polarity. 3. WNT/Ca²⁺ Pathway: This pathway involves the regula on of intracellular calcium levels and aﬀects various calcium-dependent processes within the cell. Key Genes Involved in the WNT Pathway: 1. WNT2 (Wingless-Type MMTV Integra on Site Family, Member 2): o Role: WNT2 is a member of the WNT family of signaling proteins and plays a cri cal role in the canonical WNT/β-catenin pathway. It is involved in regula ng embryonic development, cell prolifera on, and ssue regenera on. o Func on: WNT2 binds to Frizzled receptors on the cell surface, ini a ng the signaling cascade that leads to β-catenin stabiliza on and ac va on of target gene transcrip on. 2. TLE3 (Transducin-Like Enhancer of Split 3): o Role: TLE3 is a transcrip onal corepressor that interacts with transcrip on factors such as TCF/LEF in the WNT signaling pathway. It modulates the transcrip on of WNT target genes by repressing their expression. o Func on: In the WNT pathway, TLE3 can inhibit the transcrip on of WNT-responsive genes by binding to TCF/LEF transcrip on factors, thereby preven ng the ac va on of target genes in the absence of WNT signaling. Pag. 153 a 170 3. EGF (Epidermal Growth Factor): o Role: EGF is a growth factor that binds to the EGF receptor (EGFR) and ac vates downstream signaling pathways, including the WNT pathway. EGF signaling can intersect with the WNT pathway to regulate processes such as cell prolifera on, diﬀeren a on, and survival. o Func on: While EGF is not a core component of the WNT pathway, it can inﬂuence WNT signaling by modula ng the ac vity of β-catenin and other WNT pathway components, thereby aﬀec ng the transcrip on of WNT target genes. 4. ERBB3 (Erb-B2 Receptor Tyrosine Kinase 3): o Role: ERBB3 is a member of the ERBB family of receptor tyrosine kinases and is involved in various signaling pathways, including WNT signaling. ERBB3 can interact with WNT pathway components and modulate their ac vity, inﬂuencing processes like cell growth and diﬀeren a on. o Func on: ERBB3 ac va on can lead to the ac va on of downstream signaling pathways that crosstalk with the WNT pathway, impac ng the transcrip on of WNT target genes and contribu ng to the regula on of cell fate decisions. 8.5 Myogenesis Pathway: Myogenesis is the biological process by which muscle cells (myocytes) are formed. This complex process involves the diﬀeren a on of precursor cells into mature muscle ﬁbers. Myogenesis is cri cal for muscle growth, repair, and regenera on, and is regulated by a network of genes and signaling pathways that control the prolifera on, diﬀeren a on, and fusion of myogenic precursor cells into mul nucleated myotubes, which then mature into muscle ﬁbers. Genes Involved: TPM1 (Tropomyosin 1): This gene encodes a protein that is part of the tropomyosin complex, which plays a crucial role in the regula on of muscle contrac on. TPM1 is involved in stabilizing ac n ﬁlaments in muscle cells and is essen al for proper myoﬁbril assembly during muscle development. FLNC (Filamin C): FLNC encodes a protein that crosslinks ac n ﬁlaments and is involved in the organiza on of the cytoskeleton. It plays a signiﬁcant role in maintaining the structural integrity of muscle cells and in the transmission of mechanical signals during muscle contrac on. NRAP (Nebulin-related anchoring protein): NRAP is involved in the assembly of myoﬁbrils during muscle development. It interacts with other proteins in the sarcomere, the basic unit of muscle ﬁber, to help organize the ac n ﬁlaments into a func onal contrac le apparatus. 8.6 Collagen Homeostasis Pathway: Collagen homeostasis refers to the balance between the synthesis, deposi on, and degrada on of collagen in ssues. Collagen is a primary structural protein in various connec ve ssues, and maintaining its homeostasis is essen al for ssue integrity, repair, and func on. Dysregula on of collagen homeostasis can lead to ﬁbro c diseases, where excessive collagen deposi on causes ssue s ﬀness and dysfunc on, or to connec ve ssue disorders where collagen breakdown exceeds its synthesis. Genes Involved: COL6A1 (Collagen Type VI Alpha 1 Chain): This gene encodes one of the alpha chains of type VI collagen, which is a major component of the extracellular matrix (ECM) in various ssues, including muscle. Type VI collagen is involved in maintaining the structural integrity of ssues by anchoring cells to the ECM and interac ng with other ECM components. LOXL1 (Lysyl Oxidase Like 1): LOXL1 encodes an enzyme involved in the crosslinking of collagen and elas n, which is crucial for the stability and elas city of the extracellular matrix. It plays a signiﬁcant role in maintaining the structural integrity of connec ve ssues by catalyzing the forma on of covalent crosslinks between collagen molecules. TLL1 (Tolloid Like 1): TLL1 is a metalloproteinase involved in the processing and matura on of procollagen, the precursor of collagen. It cleaves procollagen to form mature collagen, which is then assembled into ﬁbers that contribute to the structural framework of ssues. Pag. 154 a 170 8.7 Pre-mRNA Processing Pathway: Pre-mRNA processing is a crucial post-transcrip onal modiﬁca on process that occurs in the nucleus of eukaryo c cells. This pathway is essen al for conver ng the primary RNA transcript (pre-mRNA) into a mature mRNA molecule that can be translated into a protein. Pre-mRNA processing includes several key steps: capping of the 5' end, splicing to remove introns, polyadenyla on of the 3' end, and edi ng or modiﬁca on of speciﬁc nucleo des. Proper regula on of this process is vital for gene expression, as errors in pre-mRNA processing can lead to defec ve proteins and are associated with various diseases. Genes Involved: UPF3A (Up-Frameshi Suppressor 3A): UPF3A is involved in the nonsense-mediated mRNA decay (NMD) pathway, which is a quality control mechanism that degrades mRNAs containing premature stop codons. Although primarily associated with NMD, UPF3A also inﬂuences pre-mRNA processing by interac ng with components of the splicing machinery and inﬂuencing the stability of mRNAs that undergo improper splicing. SRRM1 (Serine/Arginine Repe ve Matrix 1): SRRM1 is a splicing factor that plays a cri cal role in the regula on of alterna ve splicing, a process that allows a single gene to produce mul ple protein isoforms by selec vely including or excluding certain exons from the ﬁnal mRNA. SRRM1 is part of the splicing machinery and is involved in the assembly of spliceosomes, the complexes that carry out splicing. FIP1L1 (Factor Interac ng with PAPOLA and CPSF1 Like 1): FIP1L1 is involved in the polyadenyla on of the 3' end of pre-mRNA. It is a component of the cleavage and polyadenyla on speciﬁcity factor (CPSF) complex, which adds a poly(A) tail to the pre-mRNA, a modiﬁca on crucial for mRNA stability, export from the nucleus, and transla on eﬃciency. NUP98 (Nucleoporin 98): NUP98 is a component of the nuclear pore complex, which regulates the transport of molecules between the nucleus and the cytoplasm. NUP98 also plays a role in mRNA export and may inﬂuence pre-mRNA processing by interac ng with RNA and the machinery involved in splicing and polyadenyla on, thereby coordina ng the matura on and nuclear export of mRNAs. Pag. 155 a 170 8.8 Metabolic and Mitochondrial Homeostasis Pathway: Metabolic and mitochondrial homeostasis refers to the balance and regula on of metabolic processes and the maintenance of mitochondrial func on within cells. This pathway is crucial for ensuring that energy produc on, biosynthesis, and the detoxiﬁca on of harmful byproducts are carried out eﬃciently. Mitochondria are the powerhouses of the cell, playing a central role in energy produc on through oxida ve phosphoryla on, and they are also involved in regula ng apoptosis, calcium signaling, and the genera on of reac ve oxygen species (ROS). Maintaining mitochondrial homeostasis is essen al for cellular health, as disrup ons can lead to metabolic disorders, neurodegenera ve diseases, and aging-related pathologies. Genes Involved: CPS1 (Carbamoyl-Phosphate Synthetase 1): CPS1 is a key enzyme in the urea cycle, which occurs in the mitochondria of liver cells. It catalyzes the ﬁrst step in the cycle, conver ng ammonia and bicarbonate into carbamoyl phosphate. This process is crucial for the detoxiﬁca on of ammonia, a byproduct of amino acid metabolism, and is essen al for maintaining nitrogen balance and metabolic homeostasis. BHMT (Betaine—Homocysteine Methyltransferase): BHMT is an enzyme involved in the metabolism of homocysteine, a sulfur-containing amino acid. BHMT catalyzes the conversion of homocysteine to methionine using betaine as a methyl donor. This reac on is important for maintaining the balance of homocysteine levels in the body, which, if elevated, can contribute to cardiovascular disease. BHMT also plays a role in the methionine cycle, which is connected to mitochondrial func on and cellular methyla on status, impac ng overall metabolic homeostasis. SIRT3 (Sirtuin 3): SIRT3 is a mitochondrial deacetylase enzyme that regulates the func on of various mitochondrial proteins involved in energy produc on, oxida ve stress response, and apoptosis. SIRT3 is crucial for maintaining mitochondrial func on and integrity by modula ng the ac vity of enzymes involved in the tricarboxylic acid (TCA) cycle, faty acid oxida on, and the electron transport chain. SIRT3 helps to ensure eﬃcient energy produc on and protects against oxida ve stress, thus playing a vital role in mitochondrial homeostasis and longevity. ING1 (Inhibitor of Growth Family Member 1): ING1 is a tumor suppressor protein that has roles in regula ng apoptosis, cell cycle control, and DNA repair. Although primarily studied for its role in cancer biology, ING1 also inﬂuences mitochondrial func on by aﬀec ng the expression of genes involved in mitochondrial biogenesis and metabolism. Through its regula on of mitochondrial ac vity and response to cellular stress, ING1 contributes to maintaining both metabolic and mitochondrial homeostasis. Pag. 156 a 170 8.9 Inﬂamma on Hallmark Pathways: 1. HALLMARK_INFLAMMATORY_RESPONSE: o Deﬁni on: This pathway represents the set of genes involved in the body's inﬂammatory response, which is the immune system's reac on to injury or infec on. It includes various cytokines, chemokines, and other signaling molecules that help recruit immune cells to the site of injury or infec on and ac vate them to clear the threat. 2. HALLMARK_ALLOGRAFT_REJECTION: o Deﬁni on: This pathway is associated with the immune response triggered when an allogra (a transplanted organ or ssue from a donor) is recognized as foreign by the recipient's immune system. The pathway includes genes involved in T-cell ac va on and the immune processes that lead to the rejec on of the transplanted ssue. 3. HALLMARK_TNFA_SIGNALING_VIA_NFKB: o Deﬁni on: This pathway represents the signaling cascade ini ated by Tumor Necrosis Factor-alpha (TNF-α) through the ac va on of NF-κB (nuclear factor kappa-light-chain-enhancer of ac vated B cells). TNF-α is a cytokine involved in systemic inﬂamma on, and NF-κB is a transcrip on factor that controls the expression of genes involved in immune and inﬂammatory responses. 4. HALLMARK_INTERFERON_ALPHA_RESPONSE: o Deﬁni on: This pathway includes genes ac vated in response to interferon-alpha (IFN-α), a type of cytokine that is primarily involved in an viral defense. The IFN-α response is crucial for the ac va on of immune cells and the induc on of an viral states in infected and neighboring cells. 5. HALLMARK_INTERFERON_GAMMA_RESPONSE: o Deﬁni on: This pathway encompasses the genes ac vated by interferon-gamma (IFN-γ), another cytokine that plays a cri cal role in innate and adap ve immunity against viral, bacterial, and protozoal infec ons. IFN-γ is important in ac va ng macrophages, enhancing an gen presenta on, and direc ng the immune response toward a Th1 phenotype. 6. HALLMARK_IL6_JAK_STAT3_SIGNALING: o Deﬁni on: This pathway includes genes involved in the signaling cascade ini ated by Interleukin-6 (IL-6) through the JAK-STAT3 pathway. IL-6 is a cytokine that has both pro-inﬂammatory and an - inﬂammatory roles, and it is involved in the regula on of immune responses, inﬂamma on, and hematopoiesis. 7. HALLMARK_IL2_STAT5_SIGNALING: o Deﬁni on: This pathway represents the signaling cascade ini ated by Interleukin-2 (IL-2) through the STAT5 transcrip on factor. IL-2 is essen al for T-cell prolifera on, diﬀeren a on, and survival. The IL2-STAT5 signaling pathway is cri cal for maintaining immune homeostasis and the development of regulatory T cells. 8. HALLMARK_COMPLEMENT: o Deﬁni on: The complement pathway involves a group of proteins that work together to opsonize pathogens, promote inﬂamma on, and ul mately lead to the lysis (destruc on) of invading pathogens. It is a key component of the innate immune system and plays a role in the clearance of immune complexes and apopto c cells. 9. HALLMARK_COAGULATION: o Deﬁni on: This pathway involves genes that regulate blood coagula on, a process that stops bleeding by forming clots. The coagula on cascade is ghtly linked to inﬂamma on, and dysregula on of this pathway can lead to disorders like thrombosis or bleeding disorders. Pag. 157 a 170 8.10 Damage Response Hallmark Pathways: 1. HALLMARK_P53_PATHWAY: o Deﬁni on: This pathway represents the set of genes regulated by the p53 tumor suppressor protein, which plays a cri cal role in responding to cellular stress and DNA damage. When DNA is damaged, p53 is ac vated and can lead to cell cycle arrest, allowing me for DNA repair, or it can induce apoptosis if the damage is irreparable. The p53 pathway is central to preven ng the propaga on of damaged DNA, thereby protec ng against cancer. 2. HALLMARK_APOPTOSIS: o Deﬁni on: Apoptosis is a form of programmed cell death that is essen al for maintaining ssue homeostasis and elimina ng damaged or infected cells. This pathway includes genes involved in the intrinsic (mitochondrial) and extrinsic (death receptor) pathways that lead to cell death. Apoptosis is a key process in preven ng the survival of cells with severe DNA damage, thus playing a cri cal role in cancer preven on. 3. HALLMARK_REACTIVE_OXYGEN_SPECIES_PATHWAY: o Deﬁni on: Reac ve oxygen species (ROS) are chemically reac ve molecules containing oxygen that are produced as byproducts of normal cellular metabolism, par cularly in mitochondria. This pathway involves genes that regulate the produc on and detoxiﬁca on of ROS. While low levels of ROS can act as signaling molecules, excessive ROS can cause oxida ve damage to DNA, proteins, and lipids, contribu ng to aging and various diseases, including cancer. 4. HALLMARK_DNA_REPAIR: o Deﬁni on: This pathway encompasses the genes involved in the detec on and repair of DNA damage. Eﬃcient DNA repair mechanisms are crucial for maintaining genomic stability and preven ng muta ons that could lead to cancer. This pathway includes various DNA repair processes, such as nucleo de excision repair, base excision repair, mismatch repair, and homologous recombina on. 5. HALLMARK_UV_RESPONSE_UP: o Deﬁni on: This pathway represents the genes that are upregulated in response to ultraviolet (UV) radia on. UV radia on can cause direct DNA damage, leading to the forma on of cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts. The upregula on of genes in this pathway is part of the cellular response to repair this damage and protect against the carcinogenic eﬀects of UV exposure. 6. HALLMARK_UV_RESPONSE_DN: o Deﬁni on: This pathway involves the genes that are downregulated in response to UV radia on. The downregula on of certain genes following UV exposure may be a protec ve mechanism to reduce cellular ac vi es that could exacerbate DNA damage or to modulate the cell cycle to allow me for repair processes. Pag. 158 a 170 8.11 Cell Cycle Hallmark Pathways: 1. HALLMARK_MITOTIC_SPINDLE: o Deﬁni on: The mito c spindle is a structure composed of microtubules that segregates chromosomes into daughter cells during mitosis (cell division). This pathway involves genes that regulate the assembly, stability, and func on of the mito c spindle. Proper func oning of the mito c spindle is essen al for accurate chromosome segrega on, preven ng aneuploidy (an abnormal number of chromosomes), which is a hallmark of many cancers. 2. HALLMARK_E2F_TARGETS: o Deﬁni on: The E2F family of transcrip on factors is crucial for the regula on of genes required for cell cycle progression, par cularly the transi on from the G1 phase to the S phase (where DNA replica on occurs). The E2F targets pathway includes genes that are ac vated by E2F transcrip on factors, which are essen al for DNA synthesis, cell cycle progression, and the regula on of apoptosis. Dysregula on of E2F ac vity is associated with uncontrolled cell prolifera on, a characteris c of cancer. 3. HALLMARK_G2M_CHECKPOINT: o Deﬁni on: The G2/M checkpoint is a cri cal control mechanism that ensures cells do not enter mitosis (M phase) un l DNA is fully replicated and any damage is repaired. This pathway includes genes that monitor the integrity of the genome and delay the cell cycle if errors are detected. Proper func oning of the G2/M checkpoint is essen al for maintaining genomic stability and preven ng the propaga on of damaged DNA, which could lead to cancer. 4. HALLMARK_MYC_TARGETS_V1: o Deﬁni on: The MYC gene is a proto-oncogene that encodes a transcrip on factor involved in the regula on of many cellular processes, including cell growth, prolifera on, and metabolism. The MYC Targets V1 pathway includes genes directly regulated by MYC, which are involved in promo ng cell cycle progression and biomass accumula on. Overexpression of MYC is common in many cancers and is associated with aggressive tumor growth. 5. HALLMARK_MYC_TARGETS_V2: o Deﬁni on: Similar to MYC Targets V1, this pathway involves another set of genes regulated by the MYC transcrip on factor. These targets also play roles in cell cycle regula on, metabolic processes, and ribosome biogenesis. The MYC Targets V2 pathway highlights addi onal MYC-regulated genes that contribute to the control of cell prolifera on and tumorigenesis. 8.12 ECM Hallmark Pathways: 1. HALLMARK_APICAL_JUNCTION: o Deﬁni on: The apical junc on pathway involves the components and regulatory mechanisms that govern cell-cell junc ons, speciﬁcally at the apical (top) region of epithelial cells. These junc ons include ght junc ons and adherens junc ons, which are crucial for maintaining cell polarity, adhesion, and the barrier func on of epithelial layers. Proper func oning of apical junc ons is essen al for ssue integrity and communica on between cells. 2. HALLMARK_APICAL_SURFACE: o Deﬁni on: The apical surface pathway refers to the molecular features and structural components present at the apical (top) surface of polarized epithelial cells. This pathway includes proteins involved in the forma on and maintenance of the apical membrane, which plays a cri cal role in direc onal secre on, absorp on, and cell signaling. The apical surface is crucial for the specialized func ons of epithelial cells in organs like the intes nes, kidneys, and lungs. 3. HALLMARK_EPITHELIAL_MESENCHYMAL_TRANSITION (EMT): o Deﬁni on: The epithelial-mesenchymal transi on (EMT) pathway describes the process by which epithelial cells lose their cell-cell adhesion and apical-basal polarity to become more migratory and invasive mesenchymal cells. EMT is a key process in development, wound healing, and cancer metastasis. During EMT, cells downregulate epithelial markers like E-cadherin and upregulate mesenchymal markers like N-cadherin and vimen n, leading to enhanced mobility and invasiveness. Pag. 159 a 170 8.13 Metabolism Hallmark Pathways: 1. HALLMARK_GLYCOLYSIS: o Deﬁni on: Glycolysis is the metabolic pathway that converts glucose into pyruvate, genera ng small amounts of ATP (energy) in the process. This pathway is essen al for energy produc on, especially in condi ons where oxygen is limited. Glycolysis is the ﬁrst step in cellular respira on and provides intermediates for other metabolic pathways. 2. HALLMARK_OXIDATIVE_PHOSPHORYLATION: o Deﬁni on: Oxida ve phosphoryla on is the process by which cells generate ATP through the electron transport chain (ETC) in mitochondria. This pathway involves the transfer of electrons from NADH and FADH2 to oxygen, crea ng a proton gradient that drives ATP synthesis. It is the primary source of energy in aerobic organisms. 3. HALLMARK_HYPOXIA: o Deﬁni on: The hypoxia pathway is ac vated in response to low oxygen levels in ssues. Under hypoxic condi ons, cells adapt by altering their metabolism, primarily through the stabiliza on of hypoxia-inducible factors (HIFs), which regulate genes involved in glycolysis, angiogenesis, and erythropoiesis (red blood cell produc on). 4. HALLMARK_PEROXISOME: o Deﬁni on: Peroxisomes are small organelles involved in various metabolic processes, including the breakdown of very long-chain faty acids through beta-oxida on, detoxiﬁca on of hydrogen peroxide, and biosynthesis of plasmalogens (a type of phospholipid). The peroxisome pathway includes genes involved in these metabolic ac vi es, essen al for cellular lipid metabolism and redox balance. 5. HALLMARK_FATTY_ACID_METABOLISM: o Deﬁni on: This pathway involves the breakdown and synthesis of faty acids, which are cri cal sources of energy and essen al components of cell membranes. Faty acid metabolism includes beta-oxida on (faty acid breakdown) in mitochondria and peroxisomes, as well as faty acid synthesis in the cytoplasm. 6. HALLMARK_CHOLESEROL_HOMEOSTASIS: o Deﬁni on: Cholesterol homeostasis refers to the balance of cholesterol synthesis, uptake, and eﬄux in cells. This pathway includes genes involved in cholesterol biosynthesis, transport, and conversion to other molecules like bile acids. Proper cholesterol homeostasis is crucial for maintaining membrane structure, producing steroid hormones, and preven ng atherosclerosis. 7. HALLMARK_XENOBIOTIC_METABOLISM: o Deﬁni on: Xenobio c metabolism refers to the process by which the body metabolizes and detoxiﬁes foreign substances (xenobio cs) such as drugs, pollutants, and carcinogens. This pathway includes genes encoding enzymes like cytochrome P450s, which are involved in the oxida on, reduc on, and conjuga on of xenobio cs to make them easier to excrete. 8. HALLMARK_BILE_ACID_METABOLISM: o Deﬁni on: Bile acids are produced from cholesterol in the liver and play a key role in the diges on and absorp on of dietary fats. The bile acid metabolism pathway includes genes involved in the synthesis, conjuga on, and transport of bile acids. Proper regula on of bile acid metabolism is essen al for lipid diges on and preven ng liver and intes nal diseases. 9. HALLMARK_HEME_METABOLISM: o Deﬁni on: Heme metabolism involves the synthesis and breakdown of heme, a cri cal component of hemoglobin, myoglobin, and various cytochromes. This pathway includes genes involved in the biosynthesis of heme from porphyrin precursors and its degrada on to biliverdin, which is further converted to bilirubin. Proper heme metabolism is essen al for oxygen transport, electron transfer, and cellular respira on. Pag. 160 a 170 8.14 Development Hallmark Pathways: 1. HALLMARK_ANGIOGENESIS: o Deﬁni on: Angiogenesis is the process by which new blood vessels form from exis ng ones. This pathway involves the signaling molecules and cellular processes that regulate the growth, branching, and matura on of blood vessels. Angiogenesis is crucial during development, wound healing, and in certain diseases like cancer, where the growth of new blood vessels supports tumor progression. 2. HALLMARK_ADIPOGENESIS: o Deﬁni on: Adipogenesis is the process of cell diﬀeren a on by which preadipocytes develop into mature adipocytes, or fat cells. This pathway includes genes and signaling pathways that regulate the forma on and storage of fat in the body, playing a crucial role in energy balance, metabolism, and endocrine func ons. 3. HALLMARK_MYOGENESIS: o Deﬁni on: Myogenesis refers to the forma on and development of muscle ssue, par cularly skeletal muscle. This pathway encompasses the steps involved in the diﬀeren a on of myoblasts (muscle precursor cells) into mature muscle ﬁbers, as well as the signaling molecules that regulate muscle growth, repair, and regenera on. 4. HALLMARK_SPERMATOGENESIS: o Deﬁni on: Spermatogenesis is the process by which sperm cells (spermatozoa) are produced in the male testes. This pathway includes the genes and cellular processes that guide the division and matura on of spermatogonial stem cells into fully func onal sperm, essen al for male fer lity. 5. HALLMARK_PANCREAS_BETA_CELLS: o Deﬁni on: This pathway involves the development and func on of pancrea c beta cells, which are responsible for producing and secre ng insulin in response to blood glucose levels. Proper development and maintenance of beta cells are crucial for glucose homeostasis and the preven on of diabetes. 8.15 Signaling Hallmark Pathways: 1. HALLMARK_ANDROGEN_RESPONSE: o Deﬁni on: This pathway involves the cellular response to androgens, which are male sex hormones like testosterone. The androgen receptor (AR) mediates the eﬀects of androgens by regula ng the expression of genes involved in male sexual development, prostate func on, and other physiological processes. This pathway is also signiﬁcant in prostate cancer. 2. HALLMARK_ESTROGEN_RESPONSE_EARLY: o Deﬁni on: This pathway represents the early response to estrogen signaling, which is mediated by estrogen receptors (ERs). Estrogens are female sex hormones that regulate reproduc ve func ons and inﬂuence various ssues, including the breast, uterus, and bones. The early response includes immediate gene ac va on following estrogen binding. 3. HALLMARK_ESTROGEN_RESPONSE_LATE: o Deﬁni on: The late estrogen response pathway involves the downstream eﬀects of sustained estrogen signaling. This includes the regula on of genes involved in cell prolifera on, diﬀeren a on, and survival, which are important in reproduc ve ssues and breast cancer. 4. HALLMARK_NOTCH_SIGNALING: o Deﬁni on: The Notch signaling pathway is a highly conserved cell communica on system that regulates cell fate decisions. It plays a cri cal role in development, ssue homeostasis, and stem cell maintenance. Aberra ons in Notch signaling are associated with various cancers and developmental disorders. 5. HALLMARK_WNT_BETA_CATENIN_SIGNALING: o Deﬁni on: This pathway involves the Wnt signaling pathway, which regulates gene transcrip on through the stabiliza on of beta-catenin. It is crucial for embryonic development, cell prolifera on, and diﬀeren a on. Dysregula on of Wnt/beta-catenin signaling is linked to cancer, par cularly colorectal cancer. Pag. 161 a 170 6. HALLMARK_TGF_BETA_SIGNALING: o Deﬁni on: The Transforming Growth Factor-beta (TGF-β) signaling pathway regulates numerous cellular processes, including cell growth, diﬀeren a on, apoptosis, and immune responses. TGF-β signaling has dual roles in cancer, ac ng as a tumor suppressor in early stages and promo ng metastasis in advanced stages. 7. HALLMARK_HEDGEHOG_SIGNALING: o Deﬁni on: The Hedgehog signaling pathway is essen al for embryonic development, inﬂuencing cell growth, paterning, and diﬀeren a on. In adults, it maintains stem cell popula ons. Abnormal ac va on of Hedgehog signaling is implicated in several cancers, including basal cell carcinoma. 8. HALLMARK_MTORC1_SIGNALING: o Deﬁni on: The mTORC1 (mechanis c target of rapamycin complex 1) pathway is a central regulator of cell growth, prolifera on, and metabolism in response to nutrients, growth factors, and energy status. Dysregula on of mTORC1 signaling is associated with cancer, obesity, and diabetes. 9. HALLMARK_PI3K_AKT_MTOR_SIGNALING: o Deﬁni on: This pathway integrates signals from growth factors and other extracellular s muli to regulate cell survival, growth, and metabolism through the PI3K/AKT/mTOR axis. It is one of the most commonly dysregulated pathways in cancer, promo ng tumor growth and survival. 10. HALLMARK_KRAS_SIGNALING_UP: o Deﬁni on: This pathway involves the upregula on of signaling through KRAS, a small GTPase that regulates cell prolifera on, diﬀeren a on, and survival. Muta ons in KRAS are common in many cancers, leading to uncontrolled cell growth and tumorigenesis. 11. HALLMARK_KRAS_SIGNALING_DN: o Deﬁni on: This pathway represents the downregula on of KRAS signaling. Proper regula on of KRAS ac vity is essen al for normal cellular func on, and its dysregula on can lead to cancer. 12. HALLMARK_UNFOLDED_PROTEIN_RESPONSE: o Deﬁni on: The unfolded protein response (UPR) is a cellular stress response related to the endoplasmic re culum (ER). It is ac vated in response to an accumula on of unfolded or misfolded proteins in the ER, aiming to restore normal func on by enhancing protein folding capacity or degrading misfolded proteins. Chronic UPR ac va on is implicated in diseases like neurodegenera on, diabetes, and cancer. 13. HALLMARK_PROTEIN_SECRETION: o Deﬁni on: This pathway involves the processes that regulate the synthesis, folding, and traﬃcking of proteins des ned for secre on. Proper protein secre on is vital for cell communica on, immune responses, and maintaining homeostasis. Disrup ons in this pathway can lead to various diseases, including secretory disorders and cancer. 8.16 Folding, Assembly, and Pep de Loading of Class I MHC: The REACTOME gene set tled "Folding, Assembly, and Pep de Loading of Class I MHC" refers to a speciﬁc pathway involved in the prepara on of Major Histocompa bility Complex class I (MHC I) molecules for an gen presenta on on the surface of cells. Pathway Overview: This pathway describes the intricate process by which MHC class I molecules are assembled, properly folded, and loaded with pep de an gens within the endoplasmic re culum (ER) before being transported to the cell surface for presenta on to CD8+ T cells. The "Folding, Assembly, and Pep de Loading of Class I MHC" pathway is a crucial component of the immune system’s an gen presenta on mechanism, ensuring that MHC class I molecules are correctly folded, assembled, and loaded with pep des before being presented on the cell surface. This process allows the immune system to monitor cellular health and iden fy cells that are infected or transformed by pathogens or tumors, triggering appropriate immune responses. Pag. 162 a 170 Key Steps in the Pathway: 1. Synthesis and Transloca on: o MHC class I molecules are synthesized in the ribosomes and translocated into the lumen of the ER. The MHC I molecule is composed of a heavy chain (HLA class I) and a light chain (beta-2 microglobulin or B2M). 2. Chaperone-Mediated Folding: o Calnexin (CANX): Newly synthesized MHC I heavy chains ini ally bind to the ER chaperone protein calnexin, which assists in the proper folding of the heavy chain. o Calre culin (CALR): A er release from calnexin, the heavy chain associates with calre culin, another chaperone that further assists in its proper folding and stability. o ERp57 (PDIA3): This protein works alongside calre culin to ensure correct disulﬁde bond forma on within the MHC I heavy chain. 3. Associa on with Beta-2 Microglobulin (B2M): o The heavy chain then associates with B2M, which is necessary for the stability and func on of the MHC class I molecule. 4. Pep de Loading Complex (PLC): o Tapasin (TAPBP): The MHC I heavy chain-B2M complex binds to tapasin, which bridges MHC I with the Transporter associated with An gen Processing (TAP). o Transporter associated with An gen Processing (TAP1/TAP2): TAP transports pep des, which have been generated from proteasomal degrada on of cytosolic proteins, into the ER. These pep des are typically 8-10 amino acids long. o ERp57 (PDIA3): ERp57 also associates with tapasin and MHC I, aiding in the op miza on of pep de loading. o Tapasin-Related Chaperones: Addi onal components, such as tapasin and ERp57, play roles in edi ng the pep de repertoire to ensure that only high-aﬃnity pep des are loaded onto MHC I molecules. 5. Pep de Loading: o Pep des transported by TAP are loaded onto the MHC I molecules. Properly loaded pep des stabilize the MHC I complex, which is now ready to leave the ER. 6. Transport to the Cell Surface: o The stable MHC I-pep de complex is transported via the Golgi apparatus to the cell surface, where it can present the pep de an gen to CD8+ cytotoxic T cells. Importance of the Pathway: This pathway is essen al for the immune system's ability to detect and eliminate infected or malignant cells. By presen ng intracellular pep des, including those derived from viruses or mutated proteins, on the cell surface, MHC I molecules enable CD8+ T cells to recognize and destroy these compromised cells. Genes Involved: Some of the key genes involved in this pathway include: HLA-A, HLA-B, HLA-C: Encode the heavy chains of MHC class I molecules. B2M (Beta-2-Microglobulin): Encodes the light chain of MHC class I. CANX (Calnexin): Encodes the chaperone that assists in the early folding of the MHC I heavy chain. CALR (Calre culin): Encodes the chaperone that helps in the folding and stability of the MHC I complex. PDIA3 (ERp57): Encodes the protein that facilitates proper disulﬁde bond forma on and assists in pep de loading. TAP1 and TAP2: Encode the subunits of the transporter that delivers pep des from the cytosol into the ER. TAPBP (Tapasin): Encodes the bridging protein that links MHC I with TAP and aids in pep de loading. Pag. 163 a 170 8.17 Interferons: Interferons (IFNs) are a group of signaling proteins that are produced and released by host cells in response to the presence of pathogens, such as viruses, bacteria, parasites, and tumor cells. They are a cri cal part of the immune system's defense mechanism against infec ons and other threats. Interferons are named for their ability to "interfere" with viral replica on within host cells. Types of Interferons: Interferons are classiﬁed into three main types based on the type of cell that produces them and their receptor speciﬁcity: 1. Type I Interferons: o Includes: Interferon-alpha (IFN-α) and Interferon-beta (IFN-β), among others. o Produced by: Almost all cell types, par cularly in response to viral infec ons. o Func on: Type I IFNs primarily induce an an viral state in cells by upregula ng the expression of genes that inhibit viral replica on. They also ac vate natural killer (NK) cells and enhance the presenta on of an gens to T cells, boos ng the adap ve immune response. 2. Type II Interferon: o Includes: Interferon-gamma (IFN-γ). o Produced by: Immune cells, such as T cells and natural killer (NK) cells. o Func on: IFN-γ plays a crucial role in ac va ng macrophages, enhancing an gen presenta on, and promo ng the diﬀeren a on of T-helper cells towards a Th1 response. It is essen al in the defense against intracellular pathogens like bacteria and in regula ng the immune response. 3. Type III Interferons: o Includes: Interferon-lambda (IFN-λ). o Produced by: Cells of the epithelial barriers, especially in response to viral infec ons. o Func on: IFN-λ has an viral proper es similar to Type I interferons but primarily acts on epithelial cells, which are the ﬁrst line of defense against many infec ons, especially those aﬀec ng the respiratory and gastrointes nal tracts. Func ons of Interferons: An viral Response: Interferons inhibit viral replica on by inducing the expression of various an viral proteins. They help to establish an an viral state in infected and neighboring cells, making it diﬃcult for viruses to replicate and spread. Immune Ac va on: Interferons enhance the ability of immune cells to recognize and atack infected or abnormal cells. They promote the ac va on of dendri c cells, macrophages, natural killer cells, and cytotoxic T cells. Regula on of the Immune Response: Interferons play a role in modula ng the immune response to prevent overreac on, which could lead to autoimmune diseases. They help in maintaining a balance between an eﬀec ve immune response and immune tolerance. An -Tumor Ac vity: Interferons can inhibit the growth of tumors by enhancing the immune system's ability to detect and destroy cancer cells. They also directly inhibit tumor cell prolifera on and can promote the death of cancer cells. Clinical Applica ons: Interferons have been used as therapeu c agents in several diseases: Viral Infec ons: IFN-α has been used to treat chronic hepa s B and C. Cancer: IFN-α and IFN-γ are some mes used in the treatment of certain cancers, such as melanoma and some leukemias. Mul ple Sclerosis: IFN-β is used as a treatment to reduce the frequency of relapses in mul ple sclerosis. Pag. 164 a 170 8.18 Lipid Metabolism and Re nol Metabolic Pathway: Pathway Explana on: The lipid metabolism pathway is crucial for the synthesis, degrada on, and storage of lipids within the body. Lipid metabolism involves a series of biochemical processes that convert fats into energy and other essen al molecules. The re nol metabolic pathway is a speciﬁc subset of lipid metabolism, focusing on the conversion of re nol (vitamin A) into various biologically ac ve forms, which are vital for vision, immune func on, and cellular diﬀeren a on. Genes Involved: HADHB, HADH, ECH1, ECI2: These genes encode enzymes that are involved in the beta-oxida on of faty acids, a cri cal process in lipid metabolism where faty acids are broken down to generate acetyl-CoA, which then enters the citric acid cycle to produce energy. ALDH1A3, ADH1C, DHRS3: These genes are part of the re nol metabolic pathway. ALDH1A3 is involved in the oxida on of re nal to re noic acid, a key signaling molecule. ADH1C (alcohol dehydrogenase) par cipates in the oxida on of re nol to re nal. DHRS3 plays a role in the reduc on of re nal to re nol, balancing re noid levels. CYP26B1, CYP2B6: These cytochrome P450 enzymes are involved in the oxida ve metabolism of re noic acid, crucial for controlling its levels and ac vity within the body. UGT1A8: This gene encodes a UDP-glucuronosyltransferase enzyme that is involved in the glucuronida on of re noic acid, facilita ng its excre on and maintaining homeostasis. SRD5A3, GALC, BGALT5: These genes are involved in various aspects of lipid metabolism, including steroid metabolism (SRD5A3), glycolipid metabolism (GALC), and glycosyla on processes (BGALT5). 8.19 Inﬂamma on and An gen Presenta on Pathway: Pathway Explana on: The inﬂamma on and an gen presenta on pathway is cri cal for the immune system's ability to detect and respond to pathogens. Inﬂamma on is the body's immediate response to infec on or injury, while an gen presenta on involves the display of pathogen-derived pep des on cell surfaces, facilita ng the ac va on of T-cells and adap ve immune responses. Genes Involved: IRF1: Interferon-Responsive Factor 1 is a transcrip on factor that regulates genes involved in the immune response, including those that control inﬂamma on and an gen presenta on. SOCS1: Suppressor of Cytokine Signaling 1 is a nega ve regulator of cytokine signaling, playing a role in modula ng immune responses and preven ng excessive inﬂamma on. SAMD9: This gene is involved in the regula on of cell prolifera on and response to viral infec ons, contribu ng to the immune defense. TLR1: Toll-Like Receptor 1 is part of the patern recogni on receptors (PRRs) that detect pathogen- associated molecular paterns (PAMPs), triggering inﬂammatory responses. PSMB10: This gene encodes a component of the immunoproteasome, which processes intracellular proteins into pep des for presenta on on MHC class I molecules, cri cal for an gen presenta on. CTSS: Cathepsin S is involved in the processing of proteins in the lysosome, contribu ng to the degrada on of an gens for presenta on on MHC class II molecules. 8.20 Ion Channel Regula on and Epithelial Polariza on Pathway: Pathway Explana on: The ion channel regula on pathway is essen al for maintaining cellular homeostasis by controlling the ﬂow of ions across cell membranes, which is crucial for electrical signaling in neurons and muscle cells. Epithelial polariza on refers to the process by which epithelial cells organize their cellular structure, establishing dis nct apical and basal surfaces, which is vital for ssue func on and integrity. Genes Involved: KCNE3/KCNN4: These genes encode potassium channels that regulate ion ﬂow, which is essen al for maintaining the membrane poten al and electrical ac vity of cells. CYTH3: This gene is involved in regula ng ac n cytoskeleton remodeling, which is cri cal for the establishment and maintenance of epithelial cell polarity, aﬀec ng how cells interact with their environment. Pag. 165 a 170 8.21 Tumor Suppression and Membrane Phospholipid Regula on Pathway: Pathway Explana on: The tumor suppression pathway involves genes that help prevent the development and progression of cancer by regula ng cell growth, promo ng apoptosis, and maintaining genomic integrity. Membrane phospholipid regula on is cri cal for maintaining the structural integrity of cellular membranes, which is essen al for cell signaling, division, and interac on with the environment. Genes Involved: VWA5A91: This gene is a tumor suppressor that plays a role in preven ng cancerous growth by regula ng cell prolifera on and apoptosis. SAMD12/PLPP1/AGPAT3/CDS2/CPD: These genes are involved in the synthesis and remodeling of membrane phospholipids, which are vital components of the cellular membrane, aﬀec ng cell signaling, membrane ﬂuidity, and the overall func onality of the cell membrane. 8.22 Lipid Homeostasis and Tumor Suppression Pathway (Down-regulated Genes): Pathway Explana on: Lipid homeostasis is the balance between the synthesis, storage, and breakdown of lipids within the body, which is crucial for maintaining energy balance and cellular func on. The tumor suppression pathway involves the downregula on of genes that play roles in inhibi ng tumor growth and maintaining cellular structure, which might be linked to processes that enhance cell survival or diﬀeren a on. MOGAT3, DGAT2, PCK1, NR1H3, SCARB1, PCSK9, PON2, CES2, CYP2C19, CYP4F12, CYP2C18, CYP2W1, SLC16A1: These genes are involved in various aspects of lipid metabolism, including triglyceride synthesis (MOGAT3, DGAT2), faty acid oxida on (CYP4F12), cholesterol metabolism (NR1H3, PCSK9, SCARB1), and other processes crucial for maintaining lipid homeostasis. SPTLC3, DEGS2, UGT8, GAL3ST1: These genes are involved in sphingolipid metabolism, which is crucial for maintaining the structural integrity of cell membranes and signaling pathways. CDH17, ITGA6, COL17A1, DSG3, TRIM29, SERPINB5, PLOD3, LAD1, P3H2, TMEM154, S100A14, S100A16, S100A4: These down-regulated genes are involved in cell adhesion, extracellular matrix organiza on, and mo lity. The decreased expression of these genes suggests a poten al shi towards enhancing epithelial polariza on, which may contribute to the suppression of tumor progression and metastasis. 8.23 Re noid Metabolism and Epithelial Development Pathway: Pathway Explana on: The re noid metabolism and epithelial development pathway involves the regula on of genes responsible for the metabolism of re noids (vitamin A deriva ves) and the development and maintenance of epithelial ssues. Re noids are crucial for a variety of biological processes, including vision, cellular growth, diﬀeren a on, and apoptosis. They are par cularly important in the development and repair of epithelial ssues, which line organs and structures in the body. Genes Involved: CYP26B1: This gene encodes an enzyme that metabolizes re noic acid, a biologically ac ve form of vitamin A, thereby regula ng its levels within ssues and ensuring proper cellular responses. DHRS3: Dehydrogenase/Reductase 3 is involved in the reduc on of re naldehyde to re nol, which is essen al for maintaining re noid homeostasis. NRIP1: Nuclear Receptor Interac ng Protein 1 acts as a transcrip onal coregulator for nuclear receptors, including re noic acid receptors, and inﬂuences gene expression related to re noid signaling and epithelial development. RARB: Re noic Acid Receptor Beta is a nuclear receptor that mediates the biological eﬀects of re noic acid by regula ng the expression of genes involved in cell growth, diﬀeren a on, and apoptosis. GPRC5A: G Protein-Coupled Receptor Family C Group 5 Member A is a re noic acid-inducible orphan receptor that may be involved in re noid signal transduc on and epithelial cell diﬀeren a on. RARRES3: Re noic Acid Receptor Responder 3 is a gene that is induced by re noic acid and is involved in the regula on of epithelial diﬀeren a on and maintenance. APOL3: Apolipoprotein L3 is involved in lipid transport and metabolism and may also play a role in the regula on of apoptosis and inﬂamma on in epithelial cells. Pag. 166 a 170 8.24 Inﬂamma on and An gen Presenta on Pathway: Pathway Explana on: The inﬂamma on and an gen presenta on pathway is cri cal for the immune system's ability to detect and respond to pathogens and abnormal cells. Inﬂamma on is the body's immediate response to harmful s muli, such as pathogens or damaged cells, while an gen presenta on involves the process of displaying pathogen-derived pep des on the surface of cells. This allows T-cells to recognize and atack infected or cancerous cells, thereby ac va ng the adap ve immune response. Genes Involved: IRF1: Interferon Regulatory Factor 1 is a transcrip on factor that plays a central role in regula ng genes involved in the immune response, par cularly in the ac va on of genes involved in inﬂamma on and an gen presenta on. BIRC3: Baculoviral IAP Repeat Containing 3 is involved in the inhibi on of apoptosis (programmed cell death) and regula on of inﬂammatory responses. TLR3: Toll-Like Receptor 3 is a patern recogni on receptor that recognizes double-stranded RNA, a molecular patern associated with viral infec ons, and ini ates inﬂammatory responses. USF1: Upstream Transcrip on Factor 1 is involved in regula ng gene expression related to metabolism, inﬂamma on, and immune responses. CCL2: C-C Mo f Chemokine Ligand 2 is a cytokine that recruits monocytes, memory T cells, and dendri c cells to sites of inﬂamma on, playing a key role in the immune response. PSME1, PSMB10: These genes encode components of the immunoproteasome, which processes intracellular proteins into pep des that are presented on MHC class I molecules for recogni on by cytotoxic T cells. TAPBPL2: TAP Binding Protein-Like 2 is involved in the loading of an genic pep des onto MHC class I molecules, a crucial step in an gen presenta on. ERAP1, ERAP2: Endoplasmic Re culum Aminopep dase 1 and 2 are enzymes that trim pep des to the op mal length for loading onto MHC class I molecules, ensuring eﬀec ve an gen presenta on. CTSS: Cathepsin S is a protease involved in the degrada on of proteins in the lysosome, contribu ng to the processing of an gens for presenta on on MHC class II molecules. Pag. 167 a 170 8.25 Down-Regula on of ECM Pathways in ATRA-Treated Gastric Cancer Samples: In the context of gastric cancer, the downregula on of the pathway involving interac ons and signaling of the Extracellular Matrix (ECM) when samples are treated with ATRA (All-Trans Re noic Acid) compared to DMSO (Dimethyl Sulfoxide) suggests that ATRA treatment leads to a decrease in the expression and ac vity of genes and proteins associated with the ECM. The ECM is cri cal for maintaining ssue structure and providing biochemical signals that inﬂuence cell behavior, including adhesion, migra on, prolifera on, and survival. In cancer, the ECM o en becomes remodeled to facilitate tumor growth, invasion, and metastasis. Extracellular Matrix (ECM) and its Role in Cancer: The ECM consists of a network of proteins, including collagens, laminins, ﬁbronec ns, and proteoglycans, which provide structural support to cells. It also contains signaling molecules that interact with cell surface receptors like integrins, thereby inﬂuencing various cellular func ons. In cancer, the ECM undergoes signiﬁcant remodeling, which can promote tumor progression by enhancing cell migra on, invasion, and resistance to apoptosis. The ECM also plays a role in angiogenesis (the forma on of new blood vessels), which is essen al for tumor growth and metastasis. Eﬀects of ATRA on ECM Pathways: ATRA, a deriva ve of vitamin A, is known to induce diﬀeren a on and inhibit prolifera on in various cancer cells, including gastric cancer. It exerts its eﬀects by modula ng gene expression through the ac va on of re noic acid receptors (RARs), which func on as transcrip on factors. Downregula on of ECM-related genes: When gastric cancer cells are treated with ATRA, the downregula on of ECM-related pathways indicates that ATRA may reduce the expression of key ECM components and signaling molecules. This can lead to a less favorable environment for tumor cell adhesion, invasion, and metastasis. Poten al impacts: o Reduced Tumor Invasiveness: By downregula ng ECM interac ons, ATRA may impair the ability of cancer cells to migrate and invade surrounding ssues, thus limi ng the spread of the tumor. o Altered Cell-ECM Signaling: ATRA's eﬀect on ECM signaling could disrupt the communica on between cancer cells and their microenvironment, leading to reduced cell survival and prolifera on. o Decreased Angiogenesis: Since the ECM plays a role in angiogenesis, its downregula on might also impair the forma on of new blood vessels, which are necessary for tumor growth and nutrient supply. Speciﬁc Genes and Proteins Involved: While the speciﬁc genes aﬀected by ATRA in the ECM pathway can vary, common targets include: Collagens (e.g., COL1A1, COL3A1): These are structural proteins that provide tensile strength to ssues. Their downregula on can weaken the ECM, reducing its support for tumor cells. Matrix Metalloproteinases (MMPs): These enzymes degrade ECM components and facilitate ssue remodeling and invasion. ATRA may reduce MMP ac vity, thereby inhibi ng the breakdown of the ECM that is necessary for tumor invasion. Integrins: These receptors mediate cell-ECM interac ons. Downregula on of integrin expression can decrease cell adhesion and signaling, leading to reduced cell mo lity and survival. Pag. 168 a 170 8.26 Up-Regula on of Inﬂamma on/Immune Response Pathways in ATRA-Treated Gastric Cancer Samples: The up-regula on of pathways related to "Inﬂamma on, Innate Immune Response, Apoptosis, Adap ve Immune Response, An gen Presenta on, and Interferon Response" in gastric carcinoma samples treated with ATRA (All-Trans Re noic Acid) compared to the control (DMSO) suggests several poten al eﬀects on tumor behavior and the immune environment: 1. Enhanced Immune Surveillance: o Innate and Adap ve Immune Response: Up-regula on of these pathways indicates an increased ac va on of both the innate and adap ve arms of the immune system. This could enhance the recogni on and elimina on of cancer cells by immune cells, such as natural killer (NK) cells, macrophages, T cells, and B cells. 2. Increased Inﬂamma on: o Inﬂamma on Pathway: While inﬂamma on can be a double-edged sword, in this context, the up-regula on may contribute to an an -tumor immune response by recrui ng immune cells to the tumor microenvironment. However, chronic inﬂamma on can also promote tumor progression, so the overall impact may depend on the balance and dura on of the inﬂammatory response. 3. Promo on of Apoptosis: o Apoptosis Pathway: Up-regula on of apoptosis-related pathways suggests that ATRA treatment may lead to increased programmed cell death in cancer cells. This could reduce tumor cell survival and limit tumor growth. 4. Enhanced An gen Presenta on: o An gen Presenta on: Up-regula on of this pathway implies that cancer cells or surrounding immune cells may present more tumor an gens on their surface. This could improve the recogni on of cancer cells by T cells, poten ally enhancing the eﬀec veness of the immune response against the tumor. 5. Ac va on of Interferon Response: o Interferon Response: The up-regula on of interferon-related pathways suggests a heightened an viral state and an enhanced ability to ﬁght cancer. Interferons are cri cal for ac va ng immune cells, increasing an gen presenta on, and inducing an an -prolifera ve state in tumor cells. Overall Implica ons: Immune-Mediated Tumor Suppression: The up-regula on of these pathways could collec vely lead to a more hos le environment for tumor cells, promo ng their recogni on and destruc on by the immune system. This could reduce tumor growth and metastasis. Poten al for Combina on Therapies: These changes suggest that ATRA might be par cularly eﬀec ve when combined with immunotherapies, as the enhanced immune response could complement other treatments aimed at boos ng immune ac vity against the tumor. Pag. 169 a 170 8.27 Down-Regula on of Cell Cycle/Prolifera on/Division Pathways in ATRA-Treated Gastric Cancer Samples: The down-regula on of pathways related to "Cell Cycle, Cell Division, Cell Prolifera on, Cell Development, Cell Diﬀeren a on, and Cell Interac on" in gastric carcinoma samples treated with ATRA (All-Trans Re noic Acid) compared to the control (DMSO) suggests several signiﬁcant eﬀects on tumor cell behavior and growth: 1. Inhibi on of Cell Prolifera on: o Cell Cycle and Cell Division: The down-regula on of these pathways indicates a reduc on in the processes that drive cell division and progression through the cell cycle. This suggests that ATRA treatment is leading to a slower rate of tumor cell prolifera on, poten ally reducing the overall growth of the tumor. 2. Suppression of Tumor Growth: o Cell Prolifera on: A decrease in cell prolifera on pathways means that fewer cancer cells are entering the cell cycle and dividing, which directly contributes to the inhibi on of tumor growth and may result in a reduc on of tumor size over me. 3. Altered Tumor Cell Development and Diﬀeren a on: o Cell Development and Diﬀeren a on: Down-regula on of these pathways suggests that ATRA may be inducing a state where tumor cells are less capable of developing and diﬀeren a ng into more aggressive forms. This could poten ally push the cells towards a more diﬀeren ated, less malignant state, making them less capable of invasion and metastasis. 4. Reduced Tumor Cell Interac ons: o Cell Interac on: The down-regula on of cell interac on pathways implies that the communica on and adhesion between tumor cells, as well as between tumor cells and the extracellular matrix or surrounding ssues, are being disrupted. This could impair the tumor’s ability to invade neighboring ssues and metastasize. 5. Induc on of a Quiescent State: o The overall down-regula on of these pathways could lead to a state of cellular quiescence, where cancer cells remain in a non-prolifera ve state. Quiescent cells are less likely to contribute to tumor growth and may be more suscep ble to certain therapies. Overall Implica ons: Tumor Suppression: The down-regula on of these pathways suggests that ATRA is eﬀec vely suppressing the aggressive characteris cs of the gastric carcinoma cells, leading to reduced tumor growth and possibly a less invasive phenotype. Poten al for Improved Therapeu c Outcomes: By inhibi ng cell cycle progression and reducing cell prolifera on, ATRA may enhance the eﬀec veness of other cancer treatments that target prolifera ng cells or rely on slowing tumor growth. Pag. 170 a 170

PhD Thesis Notes: Insights on ATRA-Sensitivity Cluster

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