07_Cell_Cycle_2024.07.19_Student.pptx

Cell Cycle Steven J. Ontiveros, MBA, PhD  Objectives Describe and explain the significance and mechanisms of the cell cycle checkpoints. Describe the roles the mitotic spindle and checkpoint complexes play in the transition from metaphase and anaphase. Exp...

Cell Cycle Steven J. Ontiveros, MBA, PhD  Objectives Describe and explain the significance and mechanisms of the cell cycle checkpoints. Describe the roles the mitotic spindle and checkpoint complexes play in the transition from metaphase and anaphase. Explain the roles cyclins and Cdks play in regulating the progression of the cell cycle. Identify and summarize the phases of the cell cycle and the morphological changes occurring within the cell.  Overview Cell Cycle Cell reproduction: duplication of cells Genetically identical daughter cells Daughters grow and divide Progression Must be carefully regulated and coordinated Ensure progeny contain intact genomes Controlled by kinases 4 Coordinated processes Cell growth DNA replication Distribution of chromosomes Cell division Histology and Cell Biology 5 th Edition, 2020  Phases of the Cell Cycle Interphase 95% of cell cycle is spent in interphase (G1, S, G2) Chromatin is de-condensed (spaghetti bowl) Cell growth and DNA replication Phases during interphase G0 – resting phase, senescence G1 – cell grows in size (metabolically active) Prepare for S-phase S – replicates DNA (synthesis) G2 – cell grows in size Prepare for mitosis M-phase - mitosis Prophase, prometaphase, metaphase, anaphase, telophase Chromosomes are condensed (X structure) Cell duplicates Chromosomes separate Cytokinesis Cell Biology 3rd Edition 2017  Cell cycle Checkpoints Regulatory Checkpoints Prevent entry into next phase Ensures coordinated sequential events Checks and balances before proceedings Restriction G1 to S Are there appropriate growth factors in the environment DNA Damage checkpoints G1 to S, S, and G2 to M Is DNA damaged? Mitotic (Spindle Assembly) Checkpoint Mitosis (Metaphase  Anaphase) Proper chromosome attachment & Cell Biology 3rd Edition 2017 alignment  Cyclins: Regulators of the Cell Cycle Cell cycle proteins Cyclins Cyclin-dependent kinases (CDKs) Regulate the progression of the cell cycle Cyclins Accumulate and degrade in a cyclical fashion Activate Cdks CDKs Protein kinases Levels are relatively constant throughout cell cycle www.cubocube.com Regulate the cell cycle Cyclin-CDK Complexes Cyclin-Cdk Complexes Complexes trigger cell cycle events Types of Complexes Cyclin D/Cdk4,6 - Restriction point & prepare for DNA replication Cyclin E/Cdk2 – Initiation of DNA synthesis Cyclin A/Cdk2 - Initiation of DNA synthesis Cyclin B/Cdk1 – Launch mitosis Mnemonic: What happens if cyclins are not regulated appropriately...dead(b) cell Cyclin D – Cdk4,6 Cyclin E – Cdk2 Cyclin A – Cdk2 Cyclin B – Cdk1 Interphase  G1-Phase G1 Phase Cell grows Metabolically prepares for S-phase Increase in the following cyclins Cyclin D/Cdk4,6 Cyclin E/Cdk2 Cell Biology 3rd Edition 2017 Histology and Cell Biology 5 th Edition, 2020  Getting Past the Restriction Point Restriction point Growth Factors  Cyclin D synthesis  CDK4,6/CycD T Regulatory point at G1 Regulated by growth factors Rb Point of commitment for the cell cycle Stimulation Growth Factors (+) Cell bypasses the restriction point Growth Factors (-) Cell cycle stops Arrest at G0 Cyclin D Induced by growth factor (GF) stimulation Synthesized as long as GF is present Histology and Cell Biology 5th Edition, 2020 CDK4,6 Form complexes with Cyclin D (G1-Cdk) Phosphorylates protein Retinoblastoma (Rb)  Retinoblastoma Regulates Cell Proliferation by Inhibiting G1-S Transition Retinoblastoma (Rb) Tumor suppressor protein RB binds/inhibits E2F (transcription factor) Inhibits progression into cell Histology and Cell Biology 5th Edition, 2020 cycle and proliferation Inhibition of Rb RB is phosphorylated (inhibited) by: Cyclin D/Cdk4,6 Cyclin E/Cdk2 E2F Transcription factor Cell Biology 3rd Edition 2017 Stimulates expression of cyclin E & A, and CDK1  Cyclin E Cyclin E and G1-S Complexes with Cdk2 (Cyclin E/Cdk2) Transition Cyclin E/Cdk2 inhibits Rb Helps cells bypass restriction point Synergistic Effect Cyclin E/Cdk2 & Cyclin D/Cdk4,6 leads to a significant level of Rb inhibition Allows the cell to progress into S- phase Cell Biology 3rd Edition 2017 Histology and Cell Biology 5 th Edition, 2020  S-Phase S Phase DNA is replicated Centrosome duplication Increase in the following cyclins Cyclin E/Cdk2 Cyclin A/Cdk2 Cell Biology 3rd Edition 2017 S Phase and Regulation of DNA Synthesis DNA Replication Must be accurate Activated by Cyclin E/Cdk2 & Cyclin A/Cdk2 (S-Cdk) Pre-Replication Complex Preparing DNA replication sites (origins of replication) in G1 by (Cyclin E/Cdk2) [G1/S- Cdk] DNA Replication Helicase is activated by Cyclin A/Cdk2 (S- Cdk) Initiates replication p53 and DNA Damage Checkpoints p53 Transcription factor Inhibited by Mdm2 Transcription regulator for p21 Stops the cell cycle when active Mechanism Active DNA damage kinases  p53 Kinases phosphorylate (activate) p53 (stabilizes) Chk2/Chk1 Translocates into nucleus (transcription factor) p21 Campbell-Walsh-Wein Urology 12th Edition, 2021 aka CDK-interacting protein 1 inhibitor Cdk1 and Cdk2 inhibitor Activated by p53 Stops progression of the cell cycle  G2 to M-Phase Increase in the following cyclin Cyclin B/Cdk1  Onset of Mitosis: G2 to M Cyclin B/Cdk1 [M-cyclin/M-Cdk] Cyclin B/Cdk1 Function Drives G2  M transition Activates other mitotic protein kinases and Accumulates in G2 proteins involved in structural reorganization Regulation of Cyclin B/Cdk1 Prepare duplicated chromosomes Activates mitotic spindle assembly Cdk-activating kinase (CAK) Activates Cyclin B/Cdk1 by phosphorylation Wee1 Phosphorylates and inhibits M-Cdk Cyclin Stockpiles Cyclin B/Cdk1 complexes B Cdc25 Phosphatase Activates Cyclin B/Cdk1 (dephosphorylation) Potentially activated by Cyclin A/Cdk2 (S-Cdk) Positive feedback Cyclin B/Cdk1 act on Cdc25 (activate) and on Wee1 (deactivate) Mitosis  Mitosis Prophase Duplicated centrosomes migrate Nuclear envelope begins to break down Prometaphase Nuclear envelope no longer present Facilitated by Cyclin B/Cdk1 Centrosomes align at opposite poles (spindle poles) Microtubules attach to chromosomes Metaphase Chromosomes align at equator of the cell  Mitosis Anaphase Sister chromatids are pulled apart Telophase Sister chromatids arrive at spindle poles Cytokinesis Formation of the contractile ring Cell membrane pinching  Mitosis Cell Biology 3rd Edition 2017  Visualizing Cell Division Because chromatin is basophilic, histological staining allows us to identify mitotic cells Small Endometrial Esophagus Skin intestine gland Mitotic figures can typically be found in certain regions of rapidly replenishing tissues 23  Mitotic Spindles & Spindle Midzone Midzone begins to form in prophase Comprises microtubules Dynamic Instability Microtubules can be very unstable Cyclin B/Cdk1 phosphorylates microtubule- associated proteins (stabilize microtubules) Spindle consists of 3 kinds of microtubules Interpolar microtubules (highly stable) Overlapping microtubules Stabilize spindle midzone Help exert pushing forces on spindle poles Kinetochore microtubules Attach to and stabilize sister chromatids Exert pushing and pulling forces on sister chromatids Astral microtubules Cell awareness and cell location Pulling forces on the spindle poles Cell Biology 3rd Edition 2017 Metaphase to Anaphase: Mitotic Checkpoint Prometaphase Nuclear envelope breaks down Microtubules attach to kinetochores chromosomes Metaphase Chromosomes line up at midzone (middle) Pulling and pushing forces allow chromosomes to align Spindle Assembly (or Mitotic) Checkpoint: Spindle Assembly (or Mitotic) Checkpoint Complex regulates cell passage into anaphase Ensures proper attachment of microtubules to kinetochore on chromosomes  Cytokinesis Cytokinesis Development of cleavage furrow Gives rise to two daughter cells Cdk1 is inactivated (due to Cyclin B degradation) Spindle Midzone Microtubule remains from mitotic spindle Recruits proteins necessary for contractile ring formation Cell Biology 3rd Edition 2017 Contractile ring Actin and myosin II filaments Pulls plasma membrane inward  pinch off