IM [2.25] Cell Differentiation PDF 2024-2025

Cell Differentiation IM [ 2.25] Ahmed Ihab Abdelaziz MD, PhD Associate Prof. Of Molecular Medicine NewGiza University (NGU)  Developmental Genetics Objectives: ◼ Explain how a differentiated cell achieves and maintains i...

Cell Differentiation IM [ 2.25] Ahmed Ihab Abdelaziz MD, PhD Associate Prof. Of Molecular Medicine NewGiza University (NGU)  Developmental Genetics Objectives: ◼ Explain how a differentiated cell achieves and maintains its mature characteristics. ◼ Explain how we know that all genes in a differentiated cell are still capable of function. 3 Objectives: At the end of these lectures you should understand: ◼ That each specialized cell type contains specific proteins to suit its function. ◼ That this is brought about by differential gene expression. ◼ That differentiated cells contain a complete functional set of genes. ◼ That somatic cell nuclear transfer can reprogram the genome. ◼ That the genome is reprogrammed in induced pluripotent stem cells. ◼ That reprogramming can be used to produce tissues for transplantation 4  General Themes in Development ◼ Development – a series of changes in the state of a cell, tissue, organ, or organism ◼ Underlying process that gives rise to the structure and function of living organisms ◼ Developmental genetics aimed at understanding how gene expression controls this process 5 General themes ◼ Sperm and egg unite to produce a zygote ◼ That diploid cell divides and develops into the embryo ◼ Cells divide and begin to arrange themselves ◼ Each cell becomes determined – destined to become a particular cell type ◼ Commitment to become a particular type of cell occurs long before a cell actually differentiates 6 ◼ Genome is a set of genes that constitute the program of development ◼ Unicellular organisms – genome controls structure and function of the single cell ◼ Multicellular organisms– genome controls cellular features and the arrangement of cells 7 Cell adhesion ◼ Each animal cell makes its own cell adhesion molecules (CAMs) ◼ Positioning of a cell within a multicellular organism is strongly influenced by the combination of contacts it makes with other cells and the extracellular matrix 8  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Cell adhesion molecules (CAMs) Cell (c) Cell adhesion: Cell-to-cell contact conveys positional information 9 Hierarchy of transcription factors ◼ Four general phases of body formation 1. Organize body along major axes 2. Organize into smaller regions (organs, legs) 3. Cells organize to produce body parts 4. Cells change morphology and become differentiated ◼ Differential gene regulation – certain genes expressed at specific phase of development in a particular cell type ◼ Specific transcription factors are expressed at each phase of body formation 10  Hierarchy of transcription factors Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Hierarchy of transcription factors Posterior Right 1 Phase 1: 2 Phase 2: Transcription factors Transcription factors cause Dorsal determine the formation the embryo to become (ventral is of the body axes and subdivided into regions underneath) control the expression of Evidence of that have properties of Anterior transcription factors of segmentation individual segments. They phase 2. also control transcription Left factors of phase 3. Head forming 3 Phase 3: 4 Phase 4: Transcription factors cause Transcription factors cause each segment and groups cells to differentiate into of segments to develop specific cell types such as specific characteristics. skin, nerve, and muscle Limbs They also control cells. forming transcription factors of phase 4. (1): Courtesy of the National Museum of Health and Medicine, Washington, D.C.; (2): © Congenital Anomaly Research Center of Kyoto University; (3–4): Courtesy of the National Museum of Health and Medicine, Washington, D.C. 11 Phase 4 – Cell differentiation ◼ Once patterns established, cells must differentiate to carry out roles ◼ Studied in mammalian cell culture lines ◼ Differential gene expression underlies cell differentiation ◼ Stem cell characteristics ❑ Capacity to divide ❑ Daughter cells can differentiate into several cell types 12  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Stem cell Cellular division Stem cell + Red Cellular Differentiating blood division cell cell Stem cell + Red Differentiating blood cell cell 13 Stem cell categories ◼ Totipotent ❑ Ultimate stem cell is fertilized egg ❑ Can produce all adult cell types ◼ Pluripotent ❑ Embryonic stem cells (ES cells) ❑ Embryonic germ cells (EG cells) ❑ Can differentiate into almost any cell but a single cell has lost the ability to produce an entire individual 14  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Totipotent Fertilized egg is totipotent. Fertilized egg  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Totipotent Fertilized egg is totipotent. Fertilized egg Pluripotent Embryonic stem cells (ES cells) Blastocyst are pluripotent. Inner cell mass ES cells  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Totipotent Fertilized egg is totipotent. Fertilized egg Pluripotent Embryonic stem cells (ES cells) Blastocyst are pluripotent. Inner cell mass ES cells Pluripotent, multipotent, or unipotent Embryonic germ cells (EG cells) are pluripotent. Other fetal cells are multipotent Fetus or unipotent. EG cells  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Totipotent Fertilized egg is totipotent. Fertilized egg Multipotent or unipotent Adult stem cells are multipotent (bone marrow cells) or unipotent (skin cells). Pluripotent Embryonic stem Adult stem cells (ES cells) Blastocyst cells are pluripotent. Inner cell mass ES cells Pluripotent, multipotent, or unipotent Embryonic germ cells (EG cells) are pluripotent. Other fetal cells are multipotent Fetus or unipotent. EG cells ◼ Multipotent ❑ Can differentiate far fewer types of cells ❑ Hematopoietic stem cells (HSCs) ◼ Unipotent ❑ Daughter cells become only one cell type ❑ Stem cells in skin produce only skin cells 19  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Hematopoietic stem cell Bone marrow 20  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Hematopoietic stem cell Cell division Bone marrow + Hematopoietic cell Hematopoietic that will differentiate stem cell 21  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Hematopoietic stem cell Cell division Bone marrow + Hematopoietic cell Hematopoietic that will differentiate stem cell OR Myeloid Lymphoid cell cell 22  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Hematopoietic stem cell Cell division Bone marrow + Hematopoietic cell Hematopoietic that will differentiate stem cell OR Myeloid Lymphoid cell cell Basophil Monocyte Eosinophil Neutrophil Dendritic cell Red Megakaryocyte blood cell Platelets Osteoclast Macrophage 23  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Hematopoietic stem cell Cell division Bone marrow + Hematopoietic cell Hematopoietic that will differentiate stem cell OR Myeloid Lymphoid cell cell T cell B cell Natural Dendritic killer cell cell Basophil Monocyte Eosinophil Neutrophil Dendritic cell Red Megakaryocyte blood cell Osteoclast Macrophage Platelets 24 25  Davis, Weintraub, and Lasser Identified Genes That Promote Muscle Cell Differentiation ◼ What causes stem cells to differentiate into a particular cell type? ◼ Certain proteins function as “master transcription factors” ◼ Initial experimental strategy to identify genes expressed only in differentiating muscle cells ◼ Narrowed down to three genes ◼ Can any of these three genes cause non-muscle cells to differentiate into muscle cells?  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. HYPOTHESIS Muscle differentiation is induced by particular genes. KEY MATERIALS Three cloned genes had been identified that were expressed only in differentiating muscle cell lines. The researchers also had fibroblast cell lines, which do not normally differentiate into muscle cells. Experimental level Conceptual level MyoA MyoD MyoH 1 In 3 separate tubes, add each of the 3 cloned MyoA MyoD MyoH genes, designated MyoA, MyoD, and MyoH. DNA 1. Fibroblast cells 2 Add fibroblast cells to the tubes and incubate 2. CaPO4 in the presence of calcium phosphate (CaPO 4), which promotes the uptake of DNA into the cells. DNA taken Fibroblast up by cell 3 Plate the cells on solid growth media. Allow the cells to grow for 3 to 5 days. Cells will express the cloned gene. MyoD MyoA MyoH  Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Now looks like 4 Examine the cells under a microscope to determine a muscle cell if they have the morphology of differentiating muscle cells. Still looks like a fibroblast Colony labeled with myosin antibody 5 Determine if the cells are synthesizing myosin, Antibodies which is a protein that is abundantly made in muscle cells. This is done by adding a labeled antibody that recognizes myosin and determining MyoD the amounts of antibody that bind. MyoA MyoH 6 THE DATA Results from step 5: Results from step 4: Colonies labeled with antibody DNA added Microscopic morphology of cells DNA added that binds to myosin? MyoA Fibroblasts MyoA No MyoD Muscle cells MyoD Yes MyoH Fibroblasts MyoH No 7 CONCLUSION The MyoD gene encodes a protein that causes cells to differentiate into skeletal muscle cells. 8 SOURCE Davis, Robert L., Weintraub, Harold, and Lassar, Andrew B. 1987. Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell 51:987–1000. ◼ MyoD was the only one to cause fibroblasts to differentiate into muscle cells ◼ Belongs to myogenic bHLH genes ❑ Found in all vertebrates and activated during skeletal muscle development ◼ Features promoting muscle cell differentiation ❑ Basic domain binds specifically to an enhancer DNA sequence that is adjacent to genes that are expressed only in muscle cells ❑ Protein contains an activation domain that stimulates the ability of RNA polymerase to initiate transcription Thank you