10 Mitosis

• G2/ M Transition: checkpoint to make sure that DNA replication is finished

• Prophase: breakdown of nuclear envelope. Chromosomes (two sister chromatids held together by cohesion complex) condense and are released into cytoplasm. Centrosomes are moving to the poles to build spindle.

• Prometaphase: bipolar spindle is set up and capture and attach to chromosomes.

• Metaphase: alignment of chromosomes on the metaphase plate: spindles form out of highly dynamic microtubules and connect to kinetochores to place the chromosomes in the middle.

• Transition Meta/Ana: check if every chromosome is aligned on the metaphase plate

• Anaphase: sister chromatids are separated from each other with the help of shrinkage of the microtubules.

• Telophase: sister chromatids arrive near to the position of the centrosomes. reestablishment of daughter cell nuclei

• Cytokinesis: cleavage of cytoplasm by contractile ring

• M/G1 transition

Cohesin ring complexes are protein complexes, which physically hold together the sister chromatids during prophase of Mitosis

Probably are already Loaded on the Chomosomes before/ during DNA replication (late G1)

Kinetochore microtubules

• directly attach to the kinetochore region on the chromosomes

• Tension across sister kinetochores

• Separation of chromosomes and then separation of sister chromatids

Interpolation microtubules

• overlapping regions with motor proteins which bind to both microtubules

• For separation of centrosomes-> provide stability

• High forces are needed to move chromosomes through the cell

Astral microtubules

• anchor the spindles to actin structures in the surface -> stability

• Positioning of centrosomes

Dynamic structure

alpha-ß heterodimers -> build up Protokoll filaments (alpha at minus end, embedded in centrosome)

Growth at plus end

13 protofilaments build a hollow structure

Dynamic instability

• growth and shrinkage = catastrophe and rescue

• Regulation by microtubules associated proteins (MAPs)

• microtubules are polymerised from the centrosomes and are searching the entire cytoplasm to get a connection to a kinetochore

• Fast grow and shrinkage

Prometaphase

• kinetochores are built on centromeric DNA (highly repetitive-> non-coding)

• consist of many different Proteins 10-40 microtubules bind to one kinetochore

• The bound of the kinetochores is extremely tight, despite the high dynamics of the microtubules

• Equal Tension on both sides

• Tension activates kinases which signal the correct alignment

Only if all chromosomes are perfectly aligned

Triggered by cleavage (by protease) of cohesin ring -> all released at once.

Shrinkage of microtubules

Spindle moves apart along with the cell enlargement (driven by Motorproteins of interpolation kinetochores

Kinetochore microtubules get shorter

CDKs triggers the onset of mitosis if 2 inhibitory phosphorylations (Wee1, Myt1) are removed.

Active CDK pool phosphorylates various substrates which results in the chromosome condensation, nuclear envelope breakdown, centrosome separation + maturation and spindle assembly

• CDKs phosphorylate laminate filaments of nuclear envelope -> laminate looses ability of interaction

• bipolar attachement of kinetochores to spindle needed-> build up of tension

• Triggered by the cleavage of cohesin ring complexes

Regulation

• Scc1 unit gets cleaved at transition point (-> release of cohesin ring) -> mediated by seperase (protease)

• -> separation of sister chromatids

• Early phases of mitosis: seperase Inhibited by securin

• In transition, the inhibitor gets inhibited by ubiquitination (degradation) -> seperase is irreversibly activated

  means that the activation of seperase is achieved by the Anaphase promoting complex mediated degradation of Securin.

Anaphase promoting complex also has substrates later in Mitosis (Telophase): responsible for degraded cyclin b which leads to irreversible inactivation of CDK1 (needed for Mitosis exit)

The checkpoint is active in early phases of mitosisand inhibits the onset of anaphase until every chromosome is properly aligned.

Unaligned kinetochores produce constantly subcomplexes (Bub, Mad,…), which diffuse through cell and inhibit APC/C.

One unaligned kinetochore is sufficient to inhibit all APC/C.

When all kinetochores are aligned, they can no longer produce diffusable subcomplexes. -> APC/C gets active -> degrade securin -> activate protease -> Anaphase can start

Usually

Damaged

• checkpoint would not wait for all chromosomes to be aligned -> abnormal cell division: Aneuploidy (-> tumorigenesis)