cell cycle.pdf

The Cell Cycle Molecular Biology Cell cycle phases G1 – – – Between Mitosis and S phase Cells grow in size Some sensors determine when cells are too big and will push cells into SPhase S phase – DNA replication – Only occurs IF cells is going to divide G2 – Short period where cells prepare to divide...

The Cell Cycle Molecular Biology Cell cycle phases G1 – – – Between Mitosis and S phase Cells grow in size Some sensors determine when cells are too big and will push cells into SPhase S phase – DNA replication – Only occurs IF cells is going to divide G2 – Short period where cells prepare to divide – Cells prepare to divide Mitosis – Prophase – Metaphase – Anaphase – Telophase Interphase G0 How are all these transitions regulated? Combination of: – TRANSCRIPTION ACTIVATION/INHIBITION – PROTEIN ACTIVATION/INHIBITION – PROTEIN DEGRADATION Start accumulating Mitotic cyclin early but it Is inactive KEY COMPONENTS CDK – Cyclin dependent kinases – Activated / inhibited Cyclins – Different ones for each phase – Protein abundance increased through transcription and reduced through degradation Ubiquitination complexes – APC/C – SCF CDK Cyclin-DependentKinases Major drivers of the cell cycle Phosphorylate a wide variety of substrates that are different in each stage Sometimes this phosphorylation leads to activation of the substrate while other it leads to its inactivation NUCLEAR LAMINA Example of CDK substrate Nuclear Lamina is right underneath the nuclear membrane Sustains and support the nuclear membrane NUCLEAR LAMINA Example of CDK substrate Nuclear Lamina is right underneath the nuclear membrane Sustains and support the nuclear membrane NUCLEAR LAMINA Example of CDK substrate Nuclear Lamina is right underneath the nuclear membrane Sustains and support the nucleus by providing mechanical support to the nuclear membrane. It is made mostly of LaminA, a protein consisting of a long a-helix that finish in a globular domain NUCLEAR LAMINA Example of CDK substrate Nuclear Lamina is right underneath the nuclear membrane Sustains and support the nucleus by providing mechanical support to the nuclear membrane. It is made mostly of LaminA, a protein consisting of a long a-helix that finish in a globular domain During mitotic prophase, CDK phosphorylates LaminA which leads to dissambly of the nuclear lamina and the corresponding membrane break down. NUCLEAR LAMINA Example of CDK substrate THE QUESTION IS : Why doesn’t CDK phosphorylate LaminA throughout the cell cycle? Answer: because is tightly regulated to only do so during Prophase CDK REGULATION Regulated by: – Phosphorylation / de-phosphorylation (PHOSPHATES) – Type of cyclin Conformational change increases the affinity for CAK CYCLINS Cyclins provide substrate specificity to CDK by binding both CDK and the subtrates Different cyclins bind different substrates and therefore regulate what is phosphorylated by CDK in each phase Conclusion: CDK only phosphorylates a target substrate when is activated and bound to a cycling that recognizes that target Cyclins Cyclins are tightly regulated during cell cycle. Different cyclins are only expressed in certain periods and rapidly degraded in other phases Cyclins According to this scheme in G2, CDK will not phosphorylate a target recognized by cyclinE because cyclin E is not there. Who regulates the regulators? As we have seen, cyclins expression are highly regulated during the cell cycle so they only appear in certain stages. To make sure that they disappear in the next stage, cyclins are often polyubiquitinated and degraded Example: mitotic cylins are transcribed during G2-M transtion and degraded during anaphase. Thus, CDK-mitotic cyclin (also known as Mitosis promoting factor) is only active from G2-M to anaphase Who regulates the regulators? Sometimes, Cyclin-CDK combinations remain inactive by the presence of inhibitors that are rapidly degraded at the proper time Example: – in yeast S phase cyclins are present in G1 but they are inactivated due to the presence of Sic1 inhibitor. – When cells commit to pass START, Sic1 is phosphorylated by active G1 CDK. This phosphorylation is followed by polyubiquitiantion and degradation – Once Sic1 is not present S-phase CDK can trigger DNA replication E3 ligase complexes and the cell cycle Two main E3 ligase complexes regulate the cell cycle: – SCF: G1-S phase and G2-M transitions – APC/C: Metaphaseanaphase transition and anaphase to telophase transition APC/C Known as the Anaphase Promoting Complex / Cyclosome E3 ligase complex: polyubiquitinate proteins and marks them for degradation When bound to CDC20 it triggers MetaphaseAnaphase transition When bound to CDH1 it triggers Anaphase to telophase transitions APC/C Metaphase to Anaphase In metaphase Kinetochores connect the centromeres of each sister chromatides with the microtubules of the mitotic spindle The system is kept in tension because each sister chromatid is pulled to different poles of the spindle Metaphase to Anaphase Sister chromatids are held together by Cohesin a protein ring complex that loads onto chromosomes before Replication and hence, helds both copies together after replication APC/C During metaphase, APC/C binds CDC20 which triggers polyubiquitination of Securin, the inhibitor of Separase. Once freed from its inhibitor, Separase, (a protease), will cleave Cohesion complex and free each sister chromatid from each other. APC/C Once chromatids are separated mitosis starts to unravel. At this point APC/C also binds CDH1 and phosphorylate a wide variety of targets that will trigger telophase and cytokinesis. The most important target of APC/CCDH1 are mitotic cyclins. Their polyubiquitination and subsequent degradation will render CDK inactive and trigger exit from mitosis. How is APC/C regulated? During S and G2 APC/C cdc20 is inhibited by Emi1 whereas CDH1 is degraded or inactivated In prophase APC/Ccdc20 is inactivated by Mad2 / Bub3 and Bub1 whereas APC/Ccdh1 is inactivated by Rae1 and Nup98 In metaphase APC/Ccdc20 is activated by no losing its interaction with Mad2 Bub1 and Bub3 whereas APC/Ccdh1 remains inactive In anaphase APC/Ccdh1 and APC/Ccdc20 SCF Regulates key cell cycle transitions: – Promotes entry into S-phase (START) by degrading inhibitors of Sphase cyclins – Promotes entry in Mitosis by degrading APC inhibitors SCF SCF START SCF promotes START entry by degrading the inhibitors of S-phase cyclins Yeast SCF target: Sic1 Human SCF targets: RB, p21, p27 START regulation G2/M: Origin recognition complex binds replication origins but they do not trigger DNA replication because mitotic Cyclins keep other factors inactive G1: Mitotic cyclins are not present anymore and this allows the loading of DNA replication factors (cdc6, cdt1, MCM2-7) onto ORCs S: S-phase Cyclins – CDK phosphorylate and activate DNA replication factors (MCM, Cdt1..) which triggers formation of DNA replication forks. SCF and APC/C The Yin and Yang of the cell cycle SCF and APC/C regulate each other. APC SCF SCF and APC/C The Yin and Yang of the cell cycle SCF and APC/C regulate each other. – In G2-M, SCF activates APC/C by degrading EMI1, an APC/C inhibitor – In late G1, SCF inhibits APC/C by degrading CDH1 SCF Emi1 APC