Exam 4

• Cell Cycle


  Eukaryotic cell cycle has two phases: interphase and M phase (mitosis)

  
• During early stages of life, cells are constantly dividing
• G0 phase


  Inactive cell cycle that most adult cells enter after mitosis
• G0 phase


  Usually reversible; cells can re-enter cell cycle if growth factors are present
• Some cells (neurons) are post-mitotic: cannot re-enter cell cycle when exposed to growth factors
• Interphase


  1. G1 phase – cell grows, prepares for DNA replication

  2. S phase – DNA is replicated

  3. G2 phase – cell grows, prepares for mitosis

  
• M phase (mitosis and cytokinesis)


  Replicated chromosomes are organized then separated into two daughter cells

  
• Mitosis


  1. Prophase

  2. Prometaphase

  3. Metaphase

  4. Anaphase

  5. Telophase

  
• Duration of cell cycle varies between different cell types
• Mitogens


  Growth factors that activate growth factor receptors and induce cells to leave G0 and enter cell cycle

  
• Cell cycle checkpoints


  1. G1 checkpoint: G1 phase, cell only replicates DNA if growth factors are present and DNA is not damaged

  2. G2 checkpoint: G2 phase, cell only enters mitosis if DNA has been completely replicated and DNA is not damaged

  3. M checkpoint: metaphase, replicated chromosomes are segregated in anaphase only if all chromosomes are attached to mitotic spindle

  
• Cyclins

  Expressed at specific times during cell cycle, activate specific Cdks

  
• Cdks

  Constantly expressed but only active when a specific cyclin is present

  
• Early experiments with frog eggs demonstrated that cytoplasm from M phase cell can induce mitosis
• Mitosis promoting factor (MPF)


  A cyclin-dependent kinase (Cdk): non-receptor kinase that activates cell cycle proteins

  
• Cyclins


  Activate Cdks and help Cdks find target proteins to phosphorylate

  
• 1. cyclin


  Plays a critical role in regulating kinase activity of M-Cdk

  
• 1. cyclin expression starts at beginning of G2 phase; M-Cdk not activated till end of G2 phase
• Wee1 kinase


  Phosphorylates active site of M-Cdk which inhibits kinase activity

  
• Stockpile of enzymes needs to be kept inactive until cell is ready to advance to mitosis in cell cycle
• Activation of M-Cdk


  At onset of mitosis, Cdc25 phosphatase removes inhibitory phosphate to activate M-Cdk
• Cells need high amount of active M-Cdk to surpass G2 checkpoint
• Cdk inhibitors (CKIs)


  Allow cells to pause cell cycle for regulation or to adapt to change (DNA damage)

  
• Post-mitotic cells have high amounts of CKIs
• Regulating concentration of cyclins within cell is important for regulating the cell cycle

  
• Some cyclins are degraded by the proteasome
• Cell division initiation


  Cell division can initiate only if quiescent (resting) cell is pulled out of G0 and into G1 phase by extracellular signals (mitogens)

  
• Myc


  When mitogens bind to EGFRs, kinase cascade activates transcription factor (AP1) which increases Myc expression; Myc increases cyclin D

  
• Cyclin D


  G1 cyclin that activates G1-Cdk

  
• Rb protein


  Important protein that becomes phosphorylated by G1-Cdk
• Passing G1 checkpoint


  Phosphorylation of Rb allows E2F transcription factor to activate expression of G1/S and S-cyclins; cell passes G1 checkpoint and initiates DNA replication

  
• S cyclins


  Activate S-Cdks after G1 checkpoint, initiate chromosome duplication in S phase; allow cells to pass G2 checkpoint; levels elevated until mitosis
• G1/S cyclins


  Activate G1/S-Cdks in late G1, allows cell to pass G1 checkpoint, and commits cell to complete cell cycle; G1/S cyclin levels fall at start of S phase

  
• S phase regulation


  1. Inactive helicases are recruited to origins of replication by Cdc6 and origin recognition complex (ORC) during G1

  2. S-Cdk phosphorylates proteins to activate helicase at origin of replication

  3. S-Cdk phosphorylates ORC and Cdc6 so origin cannot be used; helicase cannot be recruited to origin anymore

  4. New helicases do not arrive until early G1 phase (after S phase has completed)

  5. S-Cdk prevents DNA re-replication; cell will not pass G2 checkpoint until all chromosomes are duplicated

  
• Prophase


  1. Duplicated chromosomes are condensed and kinetochore proteins are assembled on centromere

  2. Centrosomes are duplicated, mitotic spindle begins to form
• Prometaphase


  1. Nuclear envelope is broken down into vesicles

  2. Chromosomes attach to microtubules made from mitotic spindle

  
• Metaphase

  1. Organization phase: chromosomes are aligned in middle of cell

  2. Each chromosome is connected to two microtubules (one from each centrosome)

  
• Anaphase


  1. Separation phase: replicated chromosomes are pulled apart by microtubules

  2. Cohesin proteins need to be degraded to separate replicated chromosomes

  3. Microtubules attached to kinetochores need to get shorter

  
• Telophase


  1. Chromosomes decondense and arrive in opposite poles of cell

  2. Nuclear envelope reforms around around chromosomes

  3. Nuclear lamins are dephosphorylated (M cyclins have been degraded, phosphatases are activated)
• Cytokinesis


  Cytokinesis splits cytoplasm into two cells at end of mitosis