Lecture Note_Module 2 PDF

Chapter 2: An Introduction to Genes and Genomes Differences Between Prokaryotic and Eukaryotic Cells Feature Prokaryotic Cells Eukaryotic Cells No true nucleus; genetic material is located True nucleus enclosed by a nuclear Nucleus...

Chapter 2: An Introduction to Genes and Genomes Differences Between Prokaryotic and Eukaryotic Cells Feature Prokaryotic Cells Eukaryotic Cells No true nucleus; genetic material is located True nucleus enclosed by a nuclear Nucleus in the nucleoid region. membrane. Size Generally smaller (0.1 - 5.0 µm). Generally larger (10 - 100 µm). Simple structure; no membrane-bound Complex structure with membrane-bound Cell Structure organelles. organelles. DNA Circular DNA; typically one chromosome. Linear DNA; multiple chromosomes. Structure Present in most; composed of Present in plants (cellulose) and fungi Cell Wall peptidoglycan. (chitin); absent in animals. Asexual reproduction mainly through binary Asexual and sexual reproduction (mitosis Reproduction fission. and meiosis). Ribosomes Smaller (70S ribosomes). Larger (80S ribosomes). Examples Bacteria and Archaea. Animals, plants, fungi, and protists. 2.7 Differences Between Plant and Animal Cells Feature Plant Cells Animal Cells Shape Typically rectangular or box-like due to cell wall. Generally irregular or round shape. Cell Wall Present, composed of cellulose. Absent. Present; sites of photosynthesis, containing Absent; cannot perform Chloroplasts chlorophyll. photosynthesis. Large central vacuole for storage and maintaining Small and numerous vacuoles; less Vacuoles turgor pressure. prominent. Absent; plant cells have a different mechanism Centrioles Present; play a role in cell division. for cell division. Energy Glycogen is the primary storage Starch is the primary storage carbohydrate. Storage carbohydrate. Feature Plant Cells Animal Cells Present; involved in digestion and Lysosomes Rare; primarily found in lower concentrations. waste removal. Cells in muscles, skin, and organs of Examples Cells in leaves, stems, and roots of plants. animals. Comprehensive table summarizing different cell organelles, their structures, and functions: Cell Organelles: Structure and Functions Organelle Structure Function Surrounded by a double membrane Stores genetic information; controls cell Nucleus (nuclear envelope); contains chromatin activities through gene expression. (DNA and proteins). Organelle Structure Function Composed of rRNA and proteins; can be Sites of protein synthesis; translate Ribosomes free in the cytoplasm or attached to the mRNA into polypeptides. rough endoplasmic reticulum. Network of membranous tubules and sacs; Rough ER synthesizes proteins; smooth Endoplasmic rough ER has ribosomes on its surface; ER synthesizes lipids, detoxifies drugs, Reticulum (ER) smooth ER lacks ribosomes. and stores calcium ions. Modifies, sorts, and packages proteins Stacked, flattened membranous sacs Golgi Apparatus and lipids for secretion or delivery to (cisternae). other organelles. Powerhouse of the cell; site of ATP Double membrane; inner membrane Mitochondria (energy) production through cellular folded into cristae; contains its own DNA. respiration. Double membrane; contains thylakoids Site of photosynthesis; converts light Chloroplasts (stacked in grana) filled with chlorophyll. energy into chemical energy (glucose). Digests macromolecules, old cell parts, Membrane-bound vesicles containing Lysosomes and microorganisms; involved in waste digestive enzymes. disposal. Membrane-bound vesicles containing Breaks down fatty acids and detoxifies Peroxisomes enzymes that produce hydrogen peroxide. harmful substances. Large membrane-bound sacs; central Storage of substances (nutrients, waste, Vacuoles vacuole in plant cells is often large and and water); helps maintain turgor filled with fluid. pressure in plant cells. Network of protein filaments Provides structural support, maintains Cytoskeleton (microtubules, microfilaments, and cell shape, and aids in cell movement intermediate filaments). and division. Pair of cylindrical structures made of Involved in cell division; helps organize Centrioles microtubules; located in the centrosome. the mitotic spindle. Plasma Phospholipid bilayer with embedded Regulates the movement of substances Membrane proteins. in and out of the cell; protects the cell. Rigid outer layer made of cellulose (in Provides structural support and Cell Wall plants) or chitin (in fungi). protection; maintains cell shape. Organelle Structure Function Dense region within the nucleus, not Produces ribosomal RNA (rRNA) and Nucleolus surrounded by a membrane. assembles ribosomes. 2.1 Overview of Genes and Genomes Genes Definition: Genes are segments of DNA that contain the instructions for building and maintaining an organism. Each gene consists of a specific sequence of nucleotides that encodes for a functional product, which is typically a protein or an RNA molecule. Function: Genes serve as templates for the synthesis of proteins, which carry out various functions in cells. They determine inherited characteristics by coding for traits such as eye color, blood type, and susceptibility to certain diseases. Structure: A typical gene is composed of: o Exons: Coding regions that are expressed and translated into proteins. o Introns: Non-coding regions that are transcribed but removed during RNA processing. Genomes Definition: A genome is the complete set of genetic material in an organism, encompassing all of its genes as well as non-coding DNA sequences. Size and Complexity: o The genome size varies significantly among organisms, with prokaryotes generally having smaller genomes (e.g., Escherichia coli has about 4.6 million base pairs) compared to eukaryotes (e.g., humans have approximately 3 billion base pairs). o Eukaryotic genomes are more complex and contain a greater proportion of non-coding DNA, which plays roles in gene regulation and chromatin structure. Human Genome Project: An international research initiative that aimed to map the entire human genome, completed in 2003. It provided insights into the genetic basis of diseases and paved the way for advancements in personalized medicine and genetic research. 2.2 Karyotyping Definition Karyotyping is a laboratory technique that involves the examination of chromosomes in a cell. It is primarily used to determine the number and structure of chromosomes, helping in the diagnosis of genetic disorders. Significance  Clinical Diagnosis: Karyotyping is crucial for diagnosing chromosomal abnormalities, including: o Aneuploidy: The presence of an abnormal number of chromosomes (e.g., Down syndrome is caused by an extra copy of chromosome 21). o Structural Abnormalities: Such as translocations (where segments from different chromosomes are exchanged) and deletions (loss of part of a chromosome), which can lead to conditions like chronic myelogenous leukemia. Prenatal Screening: Karyotyping can be used in prenatal tests to identify potential genetic disorders in fetuses. Steps in Karyotyping 1. Cell Collection: Samples can be collected from blood, bone marrow, or amniotic fluid. 2. Cell Culture: Collected cells are cultured to encourage division. 3. Harvesting: Cells are treated with colcemid to arrest them in metaphase, when chromosomes are most visible. 4. Staining: Chromosomes are stained with dyes (e.g., Giemsa stain) to create distinct banding patterns for identification. 5. Photographing: The stained chromosomes are photographed and arranged in pairs based on size and morphology for analysis. 2.3 Chromosome Organization Structure Chromatin: DNA exists in a less condensed form known as chromatin during interphase, allowing access for transcription and replication. Histones: Histones are small, positively charged proteins that facilitate the compaction of DNA. They are key components of nucleosomes, which are the basic structural units of chromatin. Levels of Organization 1. Nucleosomes: Composed of DNA wrapped around a core of eight histone proteins. This structure resembles "beads on a string," where the DNA is the string and the nucleosomes are the beads. 2. Chromatin Types: o Euchromatin: A less condensed form of chromatin that is actively involved in transcription and gene expression.(90% of entire human genome) housekeeping genes  o Heterochromatin: A highly condensed form of chromatin that is generally transcriptionally inactive and can be further categorized into constitutive (always heterochromatic) and facultative (can become euchromatic) types. o facultative heterochromatin can transition into and out of a condensed state to accommodate gene expression, while constitutive heterochromatin is rather static and gene sparse. Specific proteins and histone modifications maintain constitutive heterochromatin. o Heterochromatin is a constituent of eukaryotic genomes with functions spanning from gene expression silencing to constraining DNA replication and repair 3. Chromosome Structure: During cell division, chromatin condenses to form visible chromosomes. Each chromosome consists of two sister chromatids joined at the centromere. 2.4 Discovery of DNA Key Discoverers 1. Friedrich Miescher (1869): Discovered "nuclein" (now known as DNA) while studying the chemical composition of white blood cells, recognizing it as a unique substance. 2. Oswald Avery (1944): Conducted experiments that demonstrated DNA is the substance responsible for heredity, transforming non-virulent bacteria into virulent ones. 3. Erwin Chargaff (1940s): Formulated Chargaff's rules, establishing that adenine (A) pairs with thymine (T) and cytosine (C) pairs with guanine (G) in DNA, leading to the base-pairing concept.  4. Rosalind Franklin (1952): Utilized X-ray diffraction techniques to capture images of DNA, providing crucial evidence for its helical structure. 5. James Watson and Francis Crick (1953): Proposed the double-helix model of DNA, incorporating findings from earlier researchers, and laid the foundation for understanding DNA replication and function. Impact on Biotechnology The discovery of DNA's structure and function has significantly impacted fields such as genetics, molecular biology, and biotechnology, leading to innovations like genetic engineering, cloning, and gene therapy. 2.5 DNA Structure Double Helix Structure: DNA consists of two strands that wind around each other, forming a double helix. Each strand is made up of nucleotides, which contain a phosphate group, a sugar (deoxyribose), and a nitrogenous base. Backbone: The sugar-phosphate backbone is formed by covalent bonds between the phosphate group of one nucleotide and the sugar of the next. Base Pairs: The nitrogenous bases (A, T, C, G) pair specifically through hydrogen bonds: o Adenine (A) pairs with Thymine (T) (two hydrogen bonds). o Cytosine (C) pairs with Guanine (G) (three hydrogen bonds). Importance of Base Pairing Base pairing is crucial for DNA replication and transcription, ensuring accurate copying of genetic information. The complementary nature of the strands allows for the formation of the double helix and facilitates the processes of DNA replication and transcription. 2.6 DNA Replication Origin of Replication  The origin of replication is a specific sequence where DNA replication begins. In prokaryotes, there is a single origin, while eukaryotes have multiple origins to facilitate rapid replication. DNA Replication Process DNA replication is a semi-conservative process, meaning each new DNA molecule contains one original strand and one newly synthesized strand. 1. Helicase: This enzyme unwinds the DNA double helix at the replication fork, separating the two strands and creating two template strands. 2. Single-Strand Binding Proteins (SSBPs): These proteins bind to the separated DNA strands, preventing them from re-annealing or forming secondary structures. 3. Topoisomerase: This enzyme relieves the tension generated ahead of the replication fork by making temporary cuts in the DNA strand, allowing it to unwind. 4. Primase: This enzyme synthesizes a short RNA primer complementary to the DNA template, providing a starting point for DNA synthesis. 5. DNA Polymerase: o Function: DNA polymerase synthesizes new DNA strands by adding nucleotides complementary to the template strand. o Types: ▪ DNA Polymerase III: The primary enzyme responsible for elongation during DNA replication. It also has 3'→5' exonuclease activity for proofreading. ▪ DNA Polymerase I: Responsible for removing RNA primers and replacing them with DNA. ▪ DNA Polymerase II: 5’3’ exonuclease activity, repair (lagging stand) 6. Ligase: This enzyme joins Okazaki fragments on the lagging strand by sealing nicks in the sugar- phosphate backbone, ensuring a continuous DNA strand. 2.7 Transcription Overview Transcription is the process by which RNA is synthesized from a DNA template. It occurs in the nucleus of eukaryotic cells and in the cytoplasm of prokaryotic cells. The primary types of RNA produced include: 1. Messenger RNA (mRNA): Carries the genetic information from DNA to the ribosome for protein synthesis. 2. Ribosomal RNA (rRNA): Forms the structural and functional core of ribosomes, where protein synthesis occurs. 3. Transfer RNA (tRNA): Delivers amino acids to the ribosome during translation, matching them with the appropriate codons on the mRNA. Difference between Coding Strand and Template Strand The strand of DNA from which mRNA is formed after transcription is known as the template strand or the antisense /minus strand. The template strand is usually directed 3' to 5' in direction. The coding strand or the sense/plus strand corresponds to the same sequence as that of the mRNA strand. https://youtu.be/DA2t5N72mgw Transcription Process 1. Initiation: RNA polymerase binds to the promoter region of the gene, unwinding a small section of the DNA double helix. 2. Elongation: RNA polymerase synthesizes the RNA strand by adding complementary ribonucleotides (A, U, C, G) based on the DNA template strand. 3. Termination: RNA polymerase continues elongation until it reaches a termination signal in the DNA sequence, at which point the RNA molecule is released. One is protein-based and the other is RNA-based. 4. Rho-dependent termination is controlled by the rho protein, which tracks along behind the polymerase on the growing mRNA chain. Near the end of the gene, the polymerase encounters a run of G nucleotides on the DNA template and it stalls. As a result, the rho protein collides with the polymerase. The interaction with rho releases the mRNA from the transcription bubble.  5. Rho-independent termination is controlled by specific sequences in the DNA template strand. As the polymerase nears the end of the gene being transcribed, it encounters a region rich in C– G nucleotides. The mRNA folds back on itself, and the complementary C–G nucleotides bind together. The result is a stable hairpin that causes the polymerase to stall as soon as it begins to transcribe a region rich in A–T nucleotides. The complementary U–A region of the mRNA transcript forms only a weak interaction with the template DNA. This, coupled with the stalled polymerase, induces enough instability for the core enzyme to break away and liberate the new mRNA transcript. Transcription Factors Transcription factors are proteins that regulate gene expression by binding to specific DNA sequences in the promoter region. They can act as enhancers or repressors, controlling the rate of transcription. 2.8 RNA Processing After transcription, eukaryotic pre-mRNA undergoes several processing steps before it becomes mature mRNA ready for translation. Steps in RNA Processing 1. 5' Capping: A modified guanine nucleotide (7-methylguanylate) is added to the 5' end of the mRNA molecule. This cap protects the mRNA from degradation and assists in ribosome binding during translation. 2. Polyadenylation: A series of adenine nucleotides (poly-A tail) is added to the 3' end of the mRNA. This tail enhances the stability of the mRNA and facilitates its export from the nucleus to the cytoplasm. 3. Splicing: Non-coding sequences (introns) are removed from the pre-mRNA, and the coding sequences (exons) are joined together. This process is carried out by a complex called the spliceosome, composed of small nuclear RNAs (snRNAs) and protein factors. 2.9 Translation Overview Translation is the process of synthesizing proteins from the mRNA template, occurring in the ribosome. Steps in Translation 1. Initiation: o The small ribosomal subunit binds to the 5' end of the mRNA and scans for the start codon (AUG), where the initiator tRNA carrying methionine binds. o The large ribosomal subunit then assembles with the small subunit to form a complete ribosome. 2. Elongation: o The ribosome moves along the mRNA, and tRNA molecules bring corresponding amino acids to the ribosome. o Peptide bonds form between adjacent amino acids, creating a polypeptide chain. 3. Termination: o The ribosome reaches a stop codon (UAA, UAG, or UGA) on the mRNA. o Release factors bind to the ribosome, prompting the release of the completed polypeptide chain and disassembly of the ribosomal complex. Codons and Anticodons Codons: Triplets of nucleotides on the mRNA that specify a particular amino acid.  Anticodons: Complementary triplets of nucleotides on tRNA that pair with the corresponding codons on mRNA, ensuring the correct amino acids are added to the growing polypeptide chain. 2.10 Gene Expression and Regulation Gene Expression Gene expression refers to the process through which the information in a gene is used to synthesize a functional product, typically a protein. The regulation of gene expression is crucial for maintaining cellular functions and responding to environmental changes. Regulation Mechanisms Gene expression can be regulated at several levels: 1. Transcriptional Regulation: o Transcription Factors: Proteins that bind to specific DNA sequences to enhance or inhibit transcription. o Enhancers and Silencers: Regulatory DNA sequences that can increase or decrease the likelihood of transcription, respectively. 2. Post-Transcriptional Regulation:  o Alternative Splicing: Different combinations of exons can be joined together to produce multiple mRNA variants from a single gene. o RNA Interference (RNAi): Small RNA molecules (miRNA and siRNA) can bind to mRNA and inhibit its translation or promote its degradation. 3. Translational Regulation: o The availability of tRNA, ribosomal proteins, and other factors can influence the efficiency of translation. 4. Post-Translational Regulation: o Protein Modifications: Chemical modifications (e.g., phosphorylation, glycosylation) can affect protein activity and stability. Lac Operon: Components and Mechanism of Action The lac operon is a classic example of gene regulation in Escherichia coli and other bacteria. It provides a model for understanding how cells control the expression of genes in response to environmental changes. The lac operon specifically regulates the metabolism of lactose, a disaccharide sugar. Importance of TATA Box, sigma factor, and Shine-Dalgarno Sequence 1. TATA Box: o Definition: The TATA box is a conserved DNA sequence located in the promoter region of many eukaryotic genes, typically around 25-30 base pairs upstream of the transcription start site. Its prokaryotic homolog is Pribnow box. o Function: It serves as a binding site for transcription factors, particularly TATA-binding protein (TBP), which is part of the larger transcription factor complex. This binding is crucial for the recruitment of RNA polymerase II to initiate transcription. o Importance: The presence of the TATA box facilitates the accurate positioning of the transcription machinery, ensuring proper gene expression. Mutations in the TATA box can lead to misregulation of gene transcription, impacting cellular functions and development. 2. The σ-factor is an essential component involved in DNA transcription. The σ-factor performs two chief functions: to direct the catalytic core of RNA Polymerase to the promoter upstream of the +1 start site of transcription, and finally to assist in the initiation of strand separation of double- helical DNA, forming the transcription "bubble." It is released once transcription initiates. 3. Shine-Dalgarno Sequence: o Definition: The Shine-Dalgarno sequence is a ribosomal binding site in prokaryotic mRNA, typically located about 6-10 nucleotides upstream of the start codon (AUG).  o Function: This sequence pairs with the 16S rRNA of the small ribosomal subunit, facilitating the initiation of translation by properly positioning the ribosome at the start codon. o Importance: The Shine-Dalgarno sequence is essential for efficient translation in bacteria. Proper recognition and binding of the ribosome ensure that proteins are synthesized accurately and efficiently, affecting cell growth and function. Variations in this sequence can influence translational efficiency and protein yield. Operon : an operon is a functioning unit of DNA containing a cluster of genes under the control of a single promoter. It is a cluster of genes that are transcribed together to give a single messenger RNA (mRNA) molecule, which therefore encodes multiple proteins Components of the Lac Operon 1. Structural Genes:  o lacZ: Encodes the enzyme β-galactosidase, which breaks down lactose into glucose and galactose. o lacY: Encodes lactose permease, a membrane protein that facilitates the uptake of lactose into the cell. o lacA: Encodes thiogalactoside transacetylase, an enzyme that detoxifies certain by- products of lactose metabolism (though its role in lactose metabolism is less crucial). 2. Regulatory Elements: o Promoter (P): The site where RNA polymerase binds to initiate transcription of the lac operon. o Operator (O): A DNA sequence that acts as the binding site for the lac repressor protein. o CAP-binding site: A regulatory sequence where the catabolite activator protein (CAP) binds to enhance transcription when glucose levels are low. 3. Regulatory Genes: o lacI: Located upstream of the operon, this gene encodes the lac repressor, a protein that can bind to the operator and prevent transcription of the operon’s genes when lactose is absent. Important Proteins 1. Lac Repressor (LacI): o The lac repressor is encoded by the lacI gene and is crucial for negative regulation of the operon. o When lactose is absent, the repressor binds to the operator, blocking RNA polymerase from transcribing the lac operon. o In the presence of lactose, an isomer of lactose called allolactose binds to the repressor, causing a conformational change that prevents it from binding to the operator. This allows transcription to proceed. 2. RNA Polymerase: o This enzyme binds to the promoter to transcribe the structural genes (lacZ, lacY, and lacA) into a single polycistronic mRNA, which is then translated into their respective proteins. 3. Catabolite Activator Protein (CAP): o CAP plays a role in positive regulation of the lac operon. o When glucose levels are low, cyclic AMP (cAMP) levels increase. cAMP binds to CAP, and the CAP-cAMP complex binds to the CAP-binding site near the promoter.  o This enhances the binding of RNA polymerase to the promoter, significantly increasing transcription of the lac operon in the presence of lactose. Mechanism of Action The lac operon is regulated by both negative and positive control mechanisms, depending on the availability of lactose and glucose. 1. Negative Regulation by Lac Repressor: o In the absence of lactose, the lac repressor binds to the operator, blocking RNA polymerase from transcribing the lac operon. o When lactose is present, allolactose (an inducer) binds to the repressor, causing it to change shape and detach from the operator. This removal of the repressor allows RNA polymerase to initiate transcription. 2. Positive Regulation by CAP: o When glucose is scarce, cAMP levels rise. The cAMP binds to CAP, which in turn binds to the CAP-binding site near the promoter. o This facilitates RNA polymerase binding, increasing the rate of transcription. However, if glucose is abundant, cAMP levels drop, and CAP does not bind to the CAP-binding site, reducing the transcription of the operon even if lactose is present. Dual Regulation: Glucose and Lactose Availability The lac operon is most active when: Lactose is present (so the repressor is inactive). Glucose is absent (so cAMP-CAP complex can form and activate transcription). When both glucose and lactose are present, glucose is preferentially utilized (this is called catabolite repression), and the lac operon remains minimally expressed until glucose is depleted. Summary of Key States: 1. No Lactose, Glucose Available: The lac repressor is bound to the operator, preventing transcription. 2. Lactose Present, Glucose Absent: Allolactose inactivates the lac repressor, cAMP binds to CAP, and CAP enhances RNA polymerase binding, leading to high transcription levels. 3. Lactose and Glucose Both Present: The repressor is inactive due to allolactose, but low cAMP levels (due to high glucose) prevent CAP from activating transcription. Hence, transcription is minimal. Importance of Lac Operon The lac operon serves as a fundamental example of gene regulation, showcasing how cells can conserve energy by producing enzymes only when needed. The dual regulation (negative by the lac repressor and positive by CAP) allows the cell to integrate signals about the availability of different sugars and prioritize the utilization of glucose over lactose. This regulatory system also plays a key role in understanding molecular genetics and biotechnology applications, such as inducible gene expression systems in recombinant DNA technology. Cyclic Adenosine Monophosphate “hunger molecule” made by e. coli when glucose is low Catabolite Repression in the Lac Operon Catabolite repression is a mechanism that enables bacteria, like E. coli, to prioritize the use of certain sugars, particularly glucose, over other energy sources such as lactose. This system ensures that the cell first consumes the most efficient energy source (glucose), and only after it is depleted does it switch to metabolizing other sugars like lactose. How Catabolite Repression Works Catabolite repression is mediated through a molecule called cyclic AMP (cAMP) and the catabolite activator protein (CAP), which also may be referred to as the cAMP receptor protein (CRP). The CAP- cAMP complex plays a key role in the positive regulation of the lac operon. Here is how this repression works: 1. When Glucose is Available: o Glucose suppresses the production of cAMP by inhibiting the enzyme adenylyl cyclase, which is responsible for converting ATP into cAMP. o In the absence of cAMP, CAP cannot bind to the CAP-binding site on the lac operon’s promoter region. This means that RNA polymerase binding to the promoter is inefficient, and transcription of the lac operon is minimal, even if lactose is present.  o Essentially, when glucose is available, the cell does not waste energy producing enzymes to metabolize other sugars like lactose because glucose is a more efficient energy source. 2. When Glucose is Absent: o When glucose levels drop, the inhibition on adenylyl cyclase is lifted, allowing cAMP levels to rise. o The cAMP molecules bind to CAP, forming the CAP-cAMP complex. This complex binds to the CAP-binding site near the promoter of the lac operon, enhancing the binding of RNA polymerase to the promoter and thus boosting transcription. o In the absence of glucose and presence of lactose (which deactivates the lac repressor), the operon is transcribed at high levels, and the enzymes needed to metabolize lactose are produced. The Role of Glucose and cAMP Glucose indirectly controls the levels of cAMP in the cell: High glucose = low cAMP → CAP cannot bind to DNA → transcription of the lac operon is repressed. Low glucose = high cAMP → CAP-cAMP binds to DNA → transcription of the lac operon is enhanced. Thus, catabolite repression ensures that E. coli uses glucose first and only turns on the machinery for lactose metabolism when glucose is depleted. This hierarchical control mechanism, known as the glucose effect, enables the bacterium to conserve energy and resources. Summary of Catabolite Repression Glucose present: Low cAMP, CAP does not bind to the CAP-binding site, so even if lactose is available, the lac operon is not strongly activated. Glucose absent: High cAMP, CAP-cAMP complex binds to the CAP-binding site, enhancing transcription of the lac operon if lactose is present. In essence, catabolite repression is a form of global control in bacterial cells that regulates the expression of many operons, including the lac operon, based on the availability of glucose. This regulation allows bacteria to efficiently manage their metabolic processes and adapt to changes in nutrient availability. The epigenome refers to the collection of chemical modifications and molecular markers that regulate gene activity and expression without altering the underlying DNA sequence. These modifications influence which genes are turned on or off in a particular cell, how strongly they are expressed, and how they respond to environmental cues. The epigenome plays a crucial role in determining cell identity, development, and response to external factors. Key Components of the Epigenome 1. DNA Methylation: o One of the most well-known epigenetic modifications is the addition of a methyl group (CH₃) to the DNA molecule, typically at cytosine bases in CpG dinucleotides. o Methylation usually represses gene expression by preventing transcription factors from accessing the DNA or recruiting proteins that compact chromatin, making it less accessible. o DNA methylation is crucial for processes like X-chromosome inactivation, genomic imprinting, and silencing transposable elements. 2. Histone Modifications: o DNA is wrapped around proteins called histones to form a structure called chromatin. Histones can undergo several types of chemical modifications, including: ▪ Acetylation (addition of acetyl groups) ▪ Methylation (addition of methyl groups) ▪ Phosphorylation (addition of phosphate groups) ▪ Ubiquitination (addition of ubiquitin proteins) o These modifications change the structure of chromatin, either loosening it to make DNA accessible for transcription (as in the case of acetylation) or compacting it to silence gene expression (as seen in certain methylation patterns). 3. Non-coding RNAs: o MicroRNAs (miRNAs) and other non-coding RNAs play a role in gene silencing and regulation at the post-transcriptional level by binding to messenger RNA (mRNA) and preventing its translation into proteins. o Long non-coding RNAs (lncRNAs) can also modify chromatin structure and gene expression by interacting with DNA or histones. 4. Chromatin Remodeling: o Specialized protein complexes can physically alter the structure of chromatin, either making it more or less accessible to the transcriptional machinery, depending on the needs of the cell. Functions of the Epigenome 1. Cell Differentiation: o During development, the epigenome ensures that specific sets of genes are activated or silenced in different cell types, allowing a single genome to give rise to various cell types like neurons, muscle cells, or liver cells.  2. Response to Environmental Factors: o The epigenome is dynamic and can be influenced by factors like diet, stress, toxins, and other environmental stimuli. These changes can affect gene expression patterns and, in some cases, may be passed down to future generations. 3. Regulation of Gene Expression: o The epigenome fine-tunes the expression of genes, determining when, where, and to what extent a gene is expressed, which is essential for normal development and function. 4. Maintenance of Genome Integrity: o Epigenetic modifications, like DNA methylation, help maintain genomic stability by silencing transposable elements (which could disrupt normal gene function) and preventing unnecessary recombination. Epigenome vs. Genome While the genome is the complete set of DNA instructions inherited from one's parents, the epigenome consists of all the chemical changes that affect gene expression. Importantly, the epigenome does not alter the DNA sequence itself but can change how genes are expressed over time. Implications for Health and Disease Cancer: Abnormal epigenetic modifications, such as inappropriate DNA methylation or histone modification patterns, can lead to uncontrolled cell growth and tumor formation. Developmental Disorders: Defects in the epigenome can cause congenital disorders due to improper gene regulation during development. Aging: Epigenetic changes accumulate over time and contribute to the aging process and the development of age-related diseases. Epigenetic Therapy: Because epigenetic changes are reversible, drugs targeting the epigenome, such as DNA methyltransferase inhibitors or histone deacetylase inhibitors, are being developed as treatments for cancer and other diseases. In conclusion, the epigenome plays a critical role in gene regulation and is central to how organisms develop, respond to their environment, and maintain health. 2.11 Mutations Definition Mutations are permanent alterations in the DNA sequence that can lead to changes in gene function. They can occur spontaneously or be induced by environmental factors such as radiation and chemicals. Types of Mutations  1. Point Mutations: A change in a single nucleotide that can result in: o Silent Mutations: No change in the amino acid sequence. o Missense Mutations: Change in a single amino acid, potentially altering protein function. o Nonsense Mutations: Introduction of a premature stop codon, leading to truncated proteins. 2. Frameshift Mutations: Insertions or deletions of nucleotides that disrupt the reading frame of the gene, often resulting in completely different protein sequences downstream. 3. Chromosomal Mutations: Large-scale alterations that affect the structure or number of chromosomes, including duplications, deletions, inversions, and translocations. Effects of Mutations Beneficial Mutations: Can confer advantages, such as antibiotic resistance in bacteria or enhanced fitness in evolving populations. Harmful Mutations: Can lead to genetic disorders or contribute to diseases such as cancer, where mutations disrupt normal cell regulation and growth. 2.12 Conclusion The study of genes and genomes is fundamental to understanding biology and advancing biotechnology. Knowledge of the molecular mechanisms governing DNA replication, transcription, translation, and gene regulation provides a foundation for developing biotechnological applications and addressing genetic disorders. Continuous research in this area holds promise for innovations in medicine, agriculture, and environmental science. Table 1: Differences Between DNA and RNA Feature DNA (Deoxyribonucleic Acid) RNA (Ribonucleic Acid) Structure Double helix Single strand Sugar Deoxyribose Ribose Adenine (A), Uracil (U), Cytosine (C), Bases Adenine (A), Thymine (T), Cytosine (C), Guanine (G) Guanine (G) Involved in protein synthesis and Function Stores genetic information gene regulation Primarily in the nucleus (eukaryotes), and nucleoid Found in nucleus, cytoplasm, and Location region (prokaryotes) ribosomes Feature DNA (Deoxyribonucleic Acid) RNA (Ribonucleic Acid) Less stable, more prone to Stability More stable due to double-stranded structure degradation Synthesized from DNA during Replication Self-replicating transcription Three main types (mRNA, rRNA, Types One main type (genomic DNA) tRNA) Table 2: Differences Among mRNA, rRNA, and tRNA Feature mRNA (Messenger RNA) rRNA (Ribosomal RNA) tRNA (Transfer RNA) Carries genetic information Forms the core component Transfers specific amino Function from DNA to ribosome for of ribosomes and catalyzes acids to the ribosome protein synthesis protein synthesis during translation Combines with proteins to Cloverleaf structure with Structure Linear strand of nucleotides form ribosomes, complex an anticodon region secondary structure Found in ribosomes (both Found in the nucleus (pre- Location free and attached to Found in the cytoplasm mRNA) and cytoplasm endoplasmic reticulum) Relatively stable but has Stable and can persist for Lifespan Generally short-lived a short lifespan during long periods translation Contains codons (triplet of Does not contain codons or Contains anticodons that Codons/Anticodons nucleotides) anticodons pair with mRNA codons Table 3: Types of DNA Type of DNA Description The complete set of DNA, including all genes and non-coding sequences, found in the Genomic DNA nucleus (eukaryotes) or nucleoid region (prokaryotes). Mitochondrial Circular DNA found in mitochondria, inherited maternally, and codes for proteins DNA essential for cellular respiration. Circular DNA found in chloroplasts of plant cells, involved in photosynthesis and Chloroplast DNA inherited maternally in plants. Type of DNA Description Small, circular DNA molecules found in prokaryotes, often carrying genes that confer Plasmid DNA advantageous traits (e.g., antibiotic resistance). DNA that constitutes the genetic material of certain viruses, which can integrate into Viral DNA the host genome or exist as episomes. Table: Differences Between A-DNA, B-DNA, and Z-DNA Feature A-DNA B-DNA Z-DNA Helix Type Right-handed helix Right-handed helix Left-handed helix Pitch per turn ~28.2 Å (2.82 nm) ~34 Å (3.4 nm) ~45.6 Å (4.56 nm) Bases per turn 11 bases per turn 10 bases per turn 12 bases per turn Helix Diameter ~23 Å (2.3 nm) ~20 Å (2.0 nm) ~18 Å (1.8 nm) Sugar C2'-endo for pyrimidines, C3'-endo (deoxyribose) C2'-endo (deoxyribose) Conformation C3'-endo for purines Glycosyl Bond Anti for pyrimidines, syn for Anti Anti Conformation purines Bases are tilted (~19°) Bases are almost Zigzag backbone (irregular) Helix Tilt relative to the helix axis perpendicular to the helix axis causing a tilt Most common form under Found in DNA regions with Found in dehydrated or Conditions physiological conditions (in high GC content or under low-water environments vivo) high salt concentrations Observed in DNA-RNA Default form in most living Associated with regulatory Biological hybrids and during organisms, responsible for regions of the genome, Significance certain types of normal DNA replication and possibly involved in gene transcription transcription regulation Deep and narrow major Flat or absent major groove, Major and Wide major groove and groove, shallow and narrow and deep minor Minor Grooves narrow minor groove wide minor groove groove Additional Notes: A-DNA: Typically forms under dehydrating conditions and is more compact than B-DNA. It is also found in DNA-RNA hybrid molecules and double-stranded RNA. B-DNA: This is the most common DNA form in physiological conditions (normal cellular environment) and the form most commonly depicted in models of the DNA double helix.  Z-DNA: Its unique left-handed helix and zigzag backbone structure give it the name "Z-DNA." It has been linked to regulatory functions and is transiently formed in some active regions of the genome. These three forms represent the structural flexibility of DNA, which can adopt different conformations depending on environmental conditions and biological requirements.