MITOSIS AND CYTOKINESIS Marie Kopecká email@example.com
In a typical eukaryotic somatic cellMITOSIS= NUCLEAR DIVISIONCYTOKINESIS = DIVISION OF THE CYTOPLASM.IMPORTANCE OF MITOSIS:EXACT SEPARATION OF REPLICATEDGENOME into two originating daughter cells→ origin of genetically identical daughter cells.Inheritance of CENTROSOME.IMPORTANCE OF CYTOKINESIS:Inheritance of corresponding part of the CYTOPLASMand CELL ORGANELLES.
Rudolf VIRCHOW (1858): „OMNIS CELLULA E CELLULA”„ALL CELLS DERIVED FROM CELLS” How ?
1590 Z. and J. Janssen: light microscope1665 R. Hook: first observation of cells1674 A. v. Leeuwenhoek: microorganisms1833 Brown: cell nucleus1838 M. J. Schleiden, T. Schwan: cell theory1839 J.E.Purkyně: cell theory1866 J. G. Mendel: element (gene)1869 F. Miescher: DNA1911 J. H.Morgan: genes in chromosomes1879 W. Flemming: chromosomes during mitosis1884 Strasburger: prophase, metaphase…1888 Waldeyer: chromosome1933 E. Ruska: first transmission electron microscope1953 J. Watson, F. Crick: structure of DNA1973 L. Hartwell: genes of the cell division cycle (cdc28)1989 P. Nurse: genes of mitotic pathway in yeast (cdc2)
Carl and Gerta CORI, the 1947 Nobel LaureatesThe origin of protein phosphorylation: PROTEINKINASES From Cohen, Nature Cell Biol 2002
The mechanisms of M-phase progression: • Protein phosphorylation protein kinases • Protein defosforylation protein phosphatases • Proteolysis (protein degradation) ubiquitination→proteasomes • Rho-GTPases
EUKARYOTIC CELL DIVISION CYCLE G1 S G2 MEach stage triggered by Cyclin dependent kinase(s)+Cyclin(s),M-phase by Cdc2/CycB=Mitotic Kinase=CDK1 -phase interphase
Nobel price for Physiology and Medicine (2001) for „Discovery of Key Regulators of Cell Division Cycle”L. Hartwell 1960→budding yeast (cdc28 gene and others).P. Nurse 1970→fission yeast (cdc2 gene of mitotic kinase).T.Hunt - biochemist →discovered cyclins
CDC28 mutant blocked at the STARTcdc2 Cdc2 mutant blocked at mitosis Nurse Hartwell
CDK1Cdc2=catalytic subunit of mitotic kinase in humans
Mitotic kinase (MPF=CDK1): Catalytic subunitjoins cyclin regulatorysubunit. It is phosporylatedat two sites. Activation ofMPF requires removal ofinhibitory phosphate by
How the mitotic kinase Cdc2/CycB (=CDK1) is activated?Inactive mitotic kinase CDK1→Activated CDK1 Mitosis → Cytokinesis - Activating kinaseMitotic kinase=CDK1Catalytic subunit Regulatory subunit Inhibitory kinase Activating phosphatase
Mitotic kinases – regulators of M-phase and its checkpoints Nigg
INTERPHASE – preparing for M-phaseG1: cell growth; preparing for S phaseS: cell growth;synthesis of nuclear DNA;synthesis of histones;sister chromatids joined by cohesins;duplication of centrosomes.G2: cell growth;preparing for M-phase.
1914 Th. Boveri: the cell consists of the cytoplasm, nucleus and the centrosome From Pederson, Nature Cell Biol 2006
Meraldi P., Nigg E., Centrosome duplication, maturation and separationFEBS Letters (2002) Polokinasa,Aurora kinasa.. Cdk1, Polokinasa, Nek2… Cdk2/Cyclin E,A
POLOKINASE mutants have aberrant mitoticspindles (e.g. monopolar or multipolar).Some cancer cells have multipolar spindles andmultiple centrosomes.From Glover et al., J.Cell Biol. 1996
G2 CHECKPOINT (G2/M) : DNA damaged? DNA replication completed?...When DNA is dama-ged or incompletelyreplicated,Checkpoint kinasesCHK1 or CHK2 switchon inhibitory pathwayto block cell cycle byinhibiting mitotickinase Cdk1/Cyclin Band phosphatasecdc25C. Whencompleted, Cdc25C isreleased to activatemitotic kinaseCdk1/Cyclin B.
Prophase: reorganisation of nucleus and cytoplasm during mitosis isinduced by activated Cdk1-cyclin complexes and other kinases thatphosphorylate numerous substrates: nuclear lamins, condensins, histonesH1, H3, MAPs, motor proteins, centrosomes, Golgi apparatus… ProphaseDisappearance ofnuclear envelope Mitotic spindle originate between separating centrosomes.Chromosomes consist of two identical sister Nucleolichromatids joined by cohesins. disappear.Condensation of chromosomes – condensins.
Prometaphase PrometaphaseEach chromatid has Microtubules joinits kinetochore in kinetochorescentromere region. Nuclear envelope disappears
Metaphase Centrosomes are at opposite poles. Mitotic apparatus consists of astral, polar and kinetochore microtubules. Chromosomes are at „metaphase plate” in the equatorial plane.Kinetochores of sisterchromatids of each chromosomejoined to microtubules directedto opposite poles of the spindle.
DIC IF1882 W. Flemming From Nurse, Cell 100, 2000 TEM
Metaphase checkpoint („spindle assembly checkpoint”)Are all chromosomes joined to mitotic spindle? NO → checkpoint activated → kinetochore signaling ⇒STOP ANAPHASE! YES →checkpoint silenced → ANAPHASE
How Anaphase is started:1. Anaphase Promoting Complex (APC).2. APC ubiquitinates and induces disruption of inhibitor „SECURIN” that inhibited protease „SEPARASE”.3. SEPARASE digests protein “COHESIN” joining sister chromatids4. Sister chromatids separate and are pulled to opposite poles of the spindle5. APC induces disruption of M-phase cyclin B→ inactivation of M-Cdk→cytokinesis
Cytokinesis in bacteria- FtsZ ring Addinal and Holland,J Mol Biol 2002
CYTOKINESIS IN ANIMAL and CYTOKINESIS IN PLANT CELLFUNGAL CELL CYTOKINESIS IN PLANT CELL CYTOKINESIS IN BACTERIA – FtsZ ring
Cytokinesis is triggered by Rho-GTPase From Narumiya and Yasuda, (2006)
Medical applications of theknowledge on the dynamics ofmicrotubules in mitosis intherapy of cancers:Microtubule inhibitors TAXOLE,VINCRISTINE, VINBLASTINE…inhibit dynamic instability ofmicrotubules → block mitosesand inhibited cells die….
Further strategy: Looking for CDK inhibitors From Meijer and Raymond, Acc.Chem Res. (2003)
Mitotic kinase Cdk1-cyclin in G2 checkpoint and Metaphase checkpoint G2 checkpoint Metaphase checkpoint Nigg
Specific termsMITOSIS The process of nuclear division.CYTOKINESIS The process of cytoplasmic division.CENTROSOME The main microtubule-organizing centre of animal cells.MITOTIC SPINDLE A highly dynamic bipolar array of microtubules that form during mitosis or meiosis and serves to move duplicated chromosomes apart.SISTER CHROMATID Duplicated chromosomes.CHECKPOINT A point where the cell division cycle can be halted until conditions are suitable for the cell to proceed to the next stage.NUCLEAR LAMINA A nuclear membrane-associated protein structure made up of lamin intermediate filament proteins.KINESIN Microtubule-based molecular motor, most often directed towards the plus end of microtubules.DYNEIN Microtubule-based molecular motor that moves towards the minus end of microtubules.GAMA-TUBULIN RING COMPLEXES Ring-like multiprotein structures implicated in microtubule nucleation.KINETOCHORE Specialized assembly of proteins that binds to a region of the chromosomes called centromere.CENTROMERE A region of eukaryotic chromosome that is attached to the mitotic spindle.