Cytoskeleton_F2024_STUDENTS.pdf

Cytosol: Cytoskeleton Contact Information Karron J. James, Ph.D. Associate Professor, Dept. of Biochemistry, Cell Biology & Genetics Office: Block B – GB6 [email protected] 484-8900, ext 1041 Office hours (in-person or on MS Teams ): Mon-Fri, by appointment Reading...

Cytosol: Cytoskeleton Contact Information Karron J. James, Ph.D. Associate Professor, Dept. of Biochemistry, Cell Biology & Genetics Office: Block B – GB6 [email protected] 484-8900, ext 1041 Office hours (in-person or on MS Teams ): Mon-Fri, by appointment Reading Lippincott's Illustrated Reviews: Cell and Molecular Biology, 3e Chapter 4: Overview, Actin (omit Regulators of the gel-sol of the cytosol), Intermediate Filaments, Microtubules Goal MCB.13. Understand the components of the cytoskeleton and how they contribute to the functions of a cell Learning Objectives Given a clinical or experimental scenario or diagram, students should be able to: MCB.13.1. Differentiate the subcellular locations and functions of cytoskeletal components and their binding proteins. MCB.13.2. Distinguish the molecular mechanisms involved in polymerization or depolymerization of cytoskeletal components.  How did we get here? Components, Nucleic acids, DNA, genes NUCLEUS: Functions Gene Transcription Product goes to CYTOSOL: Components, Functions Organelles Translation of mRNA Cytoskeletal proteins including produces Proteins Different cellular or extracellular locations go to  What is the cytoskeleton? Network of filaments found in the cytosol and the nucleus All filaments bind a target; e.g. link organelles to each other and to the plasma membrane Composed of 3 main types of filaments: Microfilaments—whole-cell movement, muscle contraction, cell shape, cytokinesis Intermediate filaments—strength Microtubules—movement of cilia and flagella, intracellular trafficking, mitotic spindle formation Fig 4.2 from Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019 Goal MCB.13. Understand the components of the cytoskeleton and how they contribute to the functions of a cell Learning Objectives Given a clinical or experimental scenario or diagram, students should be able to: MCB.13.1. Differentiate the subcellular locations and functions of cytoskeletal components and their binding proteins. MCB.13.2. Distinguish the molecular mechanisms involved in polymerization or depolymerization of cytoskeletal components. Microfilaments in different cell types Muscle cells Non-muscle cells. Examples: Epithelial cells, incl in microvilli Migrating cells, eg. fibroblasts, WBC Auditory hair cells driverlayer.com  Actin is a central component of microfilaments https://www.microscopyu.com/articles/fluorescence/filtercubes/triple/dapifitctexasred/dapifitctexasredmuntjactriple.html Actin present in all cells but different isoforms predominate in various cell types Fluorescence emission intensity from a culture of fibroblast cells stained with dyes that target the intracellular mitochondrial network (red) and cytoskeletal actin filaments (green).  G-Actin Polymerizes into Actin Filaments Polarized, dynamic G-actin monomers polymerise → F-actin microfilament G-actin + ATP→polymerization ATP hydrolysis needed for polymerization Faster at plus end than at minus end Ends may be ‘capped’ to prevent polymerization or depolymerization Fig 4.3 and 4.4 from Lippincott's Illustrated Reviews: Cell and Molecular Biology, 3e, 2024  Functions of Actin Forms structural and motility systems: Provides structural support for Fig 4.9 from Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019 plasma membranes, including protrusions (eg. microvilli) Spectrin supports plasma membrane and helps maintain cell shape Dystrophin—stability of plasma membrane in muscle and non- muscle cells http://www.78stepshealth.us/plasma-membrane/the-cytoskeleton.html Functions of Actin Restricts diffusion of organelles At the cell cortex, actin network excludes organelles Provides anchor for cell adhesion molecules Contraction, in partnership with myosin Muscle cells During telophase of mitosis, actomyosin contractile ring forms Other…  Cells migrate along extracellular matrix via actin Necessary during embryogenesis, movement of axons in response to growth factors, movement of white blood cells toward site of infection, phagocytosis Leading edge temporarily attaches to extracellular matrix Fig 4.8 from Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019  Cell migration (crawling) Actin branches through activity of ARP2/3 WASp stimulates ARP2/3 to nucleate F-actin formation Fig 16-28. Alberts. 4ed. Fig 16-90. Alberts. 4ed.  What are Thin Filaments? Name given to actin, especially in muscle cells Composed mainly of F-actin people.eku.edu Tropomyosin Troponin complex (skeletal, cardiac cells) TnI TnC TnT  Myosin Thick filament Actin-binding, motor protein Binds ATP Found in muscle, non-muscle cells. Different isoforms exist Guyton and Hall Textbook of Medical Physiology (14th ed.) Fig. 6-2  Pause for RETRIEVAL PRACTICE https://www.bookwidgets.com/pl ay/DX0HiF6O- iQAEAubM2gAAA/2E7NE28/micro filaments?teacher_id=483701221 2285440 Goal MCB.13. Understand the components of the cytoskeleton and how they contribute to the functions of a cell Learning Objectives Given a clinical or experimental scenario or diagram, students should be able to: MCB.13.1. Differentiate the subcellular locations and functions of cytoskeletal components and their binding proteins. MCB.13.2. Distinguish the molecular mechanisms involved in polymerization or depolymerization of cytoskeletal components.  Intermediate Filaments Impart mechanical strength to resist mechanical forces placed on cell Eg. Fluid sheer stress in blood vessels; stress in contracting skeletal muscles No ATP nor GTP required for polymerization www.accessexcellence.org No polarity Major IFs: keratins, vimentins (incl. desmin), neurofilaments, lamins, nestin  Epithelial IFs form strong attachment sites at cell surface http://163.178.103.176/Fisiologia/general/celulas/Membrane%20Structure%20and%20Function.htm Keratins are IFs found in cytosol of epithelial cells Interconnect desmosomes in neighbouring cells to help stabilize epithelial sheets Desmosome: adhesive junction that helps keep adjacent cells joined together Associated with strong adhesion between epithelial cells and underlying extracellular matrix  Epidermolysis Bullosa simplex https://www.aad.org/public/diseases/a-z/epidermolysis-bullosa-overview https://www.nhs.uk/conditions/epidermolysis-bullosa/ https://www.nhs.uk/conditions/epidermolysis-bullosa/ Neurofilaments http://www.nature.com/scitable/content/neurons-have-elaborate-cytoskeletal-structures-14673315 Neurofilaments in nerve cell axon serve as structural support to resist breakage Extend along length of axon Microtubules (green), intermediate filaments (purple) and actin filaments (red). Amyotropic Lateral Sclerosis (ALS) https://pharmaceutical-journal.com/article/ld/what-is-amyotrophic-lateral-sclerosis Vimentin, Desmin Vimentin family of proteins includes desmin and vimentin Vimentin found in mesenchymal cells. Widely distributed in the body Desmin expressed in all types of muscle cells Important for stabilizing the contractile apparatus Lamins Lamins found exclusively in all nuclei: lamins A, B, C are part of nuclear lamina Line inner nuclear surface, protecting chromatin from damage courses.cit.cornell.edu due to mechanical stress Disassemble at start of mitosis Re-form lamina at end of mitosis Hutchinson- Gilford progeria syndrome https://www.youtube.com/watch?v=Po4Gfk8pRkE Nestin Expressed widely: neural progenitor cells, many other cell types Associated with cell proliferation Rhabdomyosarcoma cell line. Nestin stained green Neradil J, Veselska R. Nestin as a marker of cancer stem cells. Cancer Sci. 2015;106(7):803–811. doi:10.1111/cas.12691 Pause for RETRIEVAL PRACTICE https://www.bookwidgets.com/ play/-jMATbte- iQAEM3Rh2gAAA/RE79TRS/inter mediate- fi?teacher_id=483701221228544 0 Goal MCB.13. Understand the components of the cytoskeleton and how they contribute to the functions of a cell Learning Objectives Given a clinical or experimental scenario or diagram, students should be able to: MCB.13.1. Differentiate the subcellular locations and functions of cytoskeletal components and their binding proteins. MCB.13.2. Distinguish the molecular mechanisms involved in polymerization or depolymerization of cytoskeletal components.  Microtubules Cytosolic, hollow, polar cylinders made of α- and β-tubulin heterodimers Functions include: motility (cilia and flagella), chromosomal movements during cell division (mitosis, meiosis), transport of intracellular vesicles Can rapidly assemble and disassemble Can re-direct traffic in a cell by disassembling microtubules in one region and assembling in another Fig 4.13 from Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019  Assembly of Microtubules α- and β-tubulin subunits each have bound GTP during assembly Add to plus end For mitosis, microtubules disassemble, reassemble to form mitotic spindle, then disassemble for chromosomal separation Fig 16-44. Alberts. 6ed.  Recall your basic prior knowledge: what-when-how.com www2.estrellamountain.edu  Assembly of Microtubules Centrosome composed of 2 centrioles Centrosome complex ‘caps’ minus end during elongation of microtubule Fig 16-47. Alberts. 6ed.  Dynamic instability—Growth and breakdown of microtubules Minus end ‘capped’ at centrosome complex Tubulin subunits, each with bound GTP, add to plus end β-tubulin GTP hydrolysed to GDP Presence of GTP at + end encourages Fig. 4.14 Microtubule disassembly faster assembly Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019  Dynamic instability—Growth and shrinkage of microtubules Disassembly occurs from plus end Capping plus end (microtubule capping proteins) would prevent disassembly Fig. 4.14 Microtubule disassembly Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019  Vesicle and Organelle Transport Movement of intracellular vesicles, secretory vesicles, organelles, etc. Transport requires energy (ATP) Microtubule motor proteins (ATPases) needed Kinesin, kinesin-related proteins— anterograde transport Cytoplasmic dynein—retrograde transport Fig 4.17 from Lippincott's Illustrated Reviews: Cell and Molecular Biology, 2e, 2019  Movement of Cilia and Flagella www.abpischools.org.uk Click here for a 20 second video about cilia: https://www.youtube.com/watch?v=xQG3QHMxoTA Your Patient A 15-year-old male is brought to the physician because of shortness of breath during a sporting activity. He was diagnosed with cystic fibrosis at 3 months of age based on typical clinical manifestations and results from a sweat test. His history is significant for multiple respiratory tract infections. Movement of Cilia and Flagella—Axoneme http://csls-text.c.u-tokyo.ac.jp/active/06_03.html Cilia are found in respiratory tract lining (clear mucus), lining of oviduct (transport eggs to uterus) Flagellum allows sperm to swim Axoneme: 9+2 arrangement of m’tubules extending from basal body, anchored in plasma membrane Axonemal dynein (an ATPase) needed for motility Axoneme composed of m’tubules and other torresbioclan.pbworks.com proteins to allow ‘whipping’ of cilia and wave-like motion of flagella These m’tubules are stable; resist breakdown Recall your prior knowledge: www.studyblue.com  Dynamics in Mitosis In interphase, Centrosome is replicated M’tubules disassemble New centrosomes form poles of spindle M’tubules orient spindle for cell division M’tubules ‘growing’ from each centrosome attach chromatids to spindle poles Transports chromosomes to poles during anaphase © 2010 Nature Education www.nature.com Microtubules in clinical medicine Because proper m’tubule formation is essential for spindle formation and cell division, m’tubule inhibitors commonly used for cancer chemotherapy: Taxol—drug that binds to m’tubules and blocks disassembly (so chromosomal separation is blocked) Vincristine—drug that binds tubulin monomers, preventing their polymerization NOTE: Recalling the names of these drugs is NOT required Pause for RETRIEVAL PRACTICE https://www.bookwidgets.com/ play/6ZLQVrc-- iQAFmO532gAAA/DE7WMD3/mi crotubules- kn?teacher_id=48370122122854 40 Goal MCB.13. Understand the components of the cytoskeleton and how they contribute to the functions of a cell Learning Objectives Given a clinical or experimental scenario or diagram, students should be able to: MCB.13.1. Differentiate the subcellular locations and functions of cytoskeletal components and their binding proteins. MCB.13.2. Distinguish the molecular mechanisms involved in polymerization or depolymerization of cytoskeletal components. After studying with this slide-set, the 4.5 videos and your textbook, attempt the questions at this link BEFORE the flipped classroom to help you prepare for that session: https://forms.office.com/r/XHu 9PZLnHi To continue your retrieval practice after the flipped classroom session… …Remember to review the Reading Guide and attempt the recommended questions from the textbook.