cell division. stages of mitosis (animal cell) prophase: - chromosomes condense (replicated in s...

CELL DIVISION

Stages of mitosis (animal cell)

prophase:- chromosomes condense

(replicated in S phase)- centrosomes separate

(duplicated in S phase)

prometaphase:- nuclear envelope breaks down- MTs contact chromosomes,

spindle forms

metaphase:- chromosomes align at spindle

equator (metaphase plate)

anaphase:- sister chromatids separate- chromosomes move to poles- poles move apart

telophase:- nuclear envelope reforms- chromosomes decondense- interphase array of MTs reforms

cytokinesis:- contractile ring pinches cell in

two

Centrosome cycle (animal cell):

- centrosome (centriole) duplication begins at the start of S phase- remains as one complex until M phase

In early embryonic cells, the centrosome cycle can operate without a nucleus - egg cell extracts

Mediators of mitotic chromosome structure:- cohesins: deposited along the length of sister chromatids as the DNA is replicated - hold sisters together- condensins: coil DNA - mediate chromosome condensation

Structure of a spindle:3 classes of MTs

(polar MTs)

How does a spindle form, and how does it work to separate chromosomes?

Prophase: changes in MT dynamics- more MTs nucleated from centrosome- shorter, more dynamic MTs

Quantifying MT dynamics:- inject fluorescent tubulin- bleach with laser- measure recovery (newly formed MTs)- t1/2 = time to 50% recovery

MT dynamics: regulated by MAPs vs. catastrophins

higher catastrophe rates = shorter MTs

centrosomes incubated in Xenopus egg extracts

Spindle formation in vitro:mitotic extracts + DNA + centrosomes

- abnormal spindles form when ratio of MAPs:catastrophins is perturbed

no MAP (MTs are too short)

Centrosome separation in prophase is driven by plus-end motors (KLPs) - balanced by minus-end motors

Yeast mutants: identification and characterization of spindle motors

(-) (+)

Prometaphase: kinetochores capture MTs (mechanism of attachment??)

Forces that drive chromosomes to metaphase plate:- kinetochores pull chromosomes to poles: (-)end directed motors?- astral ejection force: (+)end directed motors on chromosome arms

Metaphase: - chromosomes continue to oscillate at metaphase plate(vertebrate cells)- MTs undergo poleward flux (function?)

- poleward flux of metaphase MTs can be measured with caged fluorescein

Dynamics of individual MTs can be measured with fluorescence speckle microscopy:- poleward flux of metaphase MTs occurs in kinetochore and overlap MTs but not in astral MTs

Anaphase A: - kinetochore MTs shorten- chromosomes move to poles

Fluorescent tubulin injections show locations of MT growth, depolymerization

Two models for kinetochore movement along MTs:

Anaphase B: - poles separate- overlap MTs lengthen

Model for motor protein activity in anaphase B

Bipolar spindles can assemble without centrosomes or chromosomes

Cytokinesis (animal cell)

What determines the position of the cleavage furrow?- signal from asters to cortex- signal from central spindle- chosen before mitosis (position of spindle from previous mitosis)

Contractile ring of actin and myosin:red = actin, green = myosin II

Cytokinesis in plants