Cell Cycle, Cell Division, and Apoptosis

What do cells do to prepare to divide

They grow by increasing size and number of organelles, generate more proteins, duplicate chromosomes → each stage is highly coordinate and regulated

What does each round of the cell cycle create

2 identical daughter cells

Duration of cell cycle among different cells

Varies greatly; single-celled yeast divide every 2 hours and mammalian liver cells divide once a year

What are the phases of cell cycle and what do they do (in general)

M phase, G1 phase, S phase, G2 phase; they coordinate cell growth, DNA replication, mitosis, and cell division

What is interphase

G1, S, and G2 phase

What is G1 phase

The gap between M and S phase

What is G2 phase

gap between S and M phase

What occurs in M phase

mitosis (nucleus divides) and then cytokinesis (cytoplasm divides)

What occurs during the gap phases

they give cells enough time to increase its size and number of organelles

What occurs during S-phase

DNA synthesis

What do duaghter cells have in similarity with the parent cell

correct number of chromosomes, they r the same size

What are early embryonic cleavages

DNA/division without Gap phases (the fertilized egg is large and has overabundance of organelles/proteins) → each division will result in smaller daughter cells

What are the 3 most prominent checkpoints in cell cycle

G1→S, G2→M, and whether M phase is ready to pull duplicated chromosomes apart

What are questions asked in the G1 checkpoint

Is environment favorable

What are questions asked in the G2 checkpoint

Is all DNA replicated? is all DNA damage repaired?

What are questions asked in the M checkpoint

Are all chromosomes properly attached to mitotic spindle

Wha't’s the ‘5th’ cell cycle phase in mammilian cells

G0, quiescence

What are options that cells can go through in G1 checkpoint

proceed to S phase?

pause? (until conditions are right)

withdraw and enter G0? (someitmes cells can reenter the cycle once conditions improve)

Withdraw permanently/terminally differentiate? (persist in G0 for the lifetime of the animal)

What controls the progression of the cell cycle

Fluctuations in cyclin-CDK activity

What are cyclin

regulatory proteins with no intrinsic kinase activity that binds and activated Cdk proteins, their levels fluctuate in the cell cycle (peak is mitosis and dip in interphase)

What is Cdk

cyclin-dependent kinase proteins, and they trigger cell cycle events when they are acticated

What does active cyclin-Cdk complex do

phosphorylates key proteins in the cell that are required to initiate cell cycle

What does cyclin help do in the cyclin-Cdk complex

helps direct Cdk to the target proteins that the Cdk phosphorylates

What does the formation of active cyclin-Cdk complexes drive

various cell-cycle events including entry into S phase or M phase

zwhat does increase in cyclin concentration do

helps form the active cyclin-Cdk complex that drives entry into M phase

Does concentration of Cdk component change

NO, only the enzymatic activity of the cyclin-Cdk complex rises and falls

What is the cyclin that acts in G2 to trigger entry into M phse

M cyclin and the complex it forms with Cdk is called M-Cdk

What are distinct cyclins and what do they do

They are S cyclins and G1/S cyclins; they bind to distinct Cdk protein late in G1 to form S-Cdk and G1/S-Cdk and trigger entry into S phase

What do G1 cyclins do

act earlier in G1 and bind to other Cdk proteins to form G1 Cdks, which help drive the cell through G1 towards S phase

What does activation of Cdk require

phosphorylation/dephosphorylation and cyclin binding

How is S cyclin-Cdk complexes regulated

the binding of p27, a Cdk inhibitior, which delays progression from G1-S, prevents Cdk from phosphorylating its targets

What happens once cells transition to S phase

they rapidly progress through cell cycle

What do mitogens do

promotes cell cycle entry by activating the synthessi of G1 and G1/S cyclins

What is the Rb and waht does it do

its the retinoblastoma protein and it binds/inhibits trnascription regulators

What is Rb inhibited by

G1-Cdk and G1/S-Cdk phosphorylation

What does p-Rb

releases the break on the cell cycle so transcription of genes occurs for entry into S phase

Where can cell-cycle control system arrest the cycle

middle of G1: damaged DNA

end of G1: unfavorable extracellular enviornment

end of S: damaged/incompletely replicated DNA

middle of G2: damaged/incompletely repiclicated DNA

end of M: chromosome improperly attached to mitotic spindle

What does damage in G1 do in the G1-to-S transition

increases levels and activity of transcription regulator p53

what does phos.-p53 do in the G1-to-S transition

activates transcription of Cdk inhibitor p21

What does p21 do in the G1-to-S transition

binds S-Cdk and G1/S-Cdk to block cell cycle entry

What is found in 50% cancer

mutations in p53

What part of cell cycle does p21 protein binding to G1/S-Cdk and S-Cdk occur

G1

Step 1 of how DNA replication occurs in the G1-to-S transition

the formation of the pre-replicative complex:

The origin recognition complex (ORC) binds origins of replications throughout the cell cycle

Cdc6 is present in G1 and binds ORC

DNA helicases are recruited

Step 2 of how DNA replication occurs in the G1-to-S transition

Formation of replication fork:

Accumulation of active S-Cdk

S-CDk activates the helicase and recruitment of DNA polymerase

S-Cdk phosphorylates Cdc6 which is targeted for degradation

How is M cyclin-Cdk regulated part 1

M cyclin accumulates in G2 to reach a maximum in early M. It binds to M-Cdk but phosphate inactivates it. M-Cdk if inactivated by phosphorylation at 2 sites (one of the kinase inhibitors is Wee1). An activating phosphatase removes the inhibitory phosphate from M-Cdk. The activated M-Cdk causes phosphorylation which induces choromosome condensation, dispersal of the nucleolus, and formation of spindle. M-Cdk phosphorylation of the nuclear lamina and pores cause them and the nuclear envelope to break down in prometaphase.

How does activated M-Cdk indirectly activate more M-Cdk

It’s a positive feedback loop. So, once activated M-Cdk phosphorylates and thereby activates more Cdk-activating phosphatase. Cdc25 phosphatase can now activate more M-Cdk by removing inhibitory phsophate groups from the Cdk subunit

How is cyclin degradation controlled

ubiquitin-mediated proteolysis

S and M cyclins are rapidly degraded and the anaphase promoting complex (APC) covalently attaches ubiquitin chains to the S and M cyclins and it tags those cyclin for degradation by the proteasome

What does loss of cyclin do

renders Cdk inactive

Where does inhibition of activating phosphatase Cdc25 block

entry to mitosis from G2 since DNA replication isn’t complete or ther'e’s DNA damage

Where does inhibition of APC/C activation delays occur

in the exit of mitosis because chromosomes aren’t properly attached to spindle

Where does Cdk inhibitors block entry

into s phase because environment not favorable

What happens at metaphase

chromsomes line up at equator

What happens at anaphase

chromatids separate

What are the fates of the daughter cells

they are often different, could be stem cell or differentiated cells

How does M-Cdk shut down PP2A-B55, a phosphatase

it inhibits PP2A-B55 that opposes its effects on downstream targets, and this inhibition allows targets of M-Cdk to drive cells into mitosis

What does each chromosome have

1 centromere, 2 telomeres, and many origins of replication

What are chromatids

duplicated chromsomes tied togehter

What is the centromere

contains repeated base sequences (heterochromatin) and is visible as a constriction of the chromosome

Whats the kinetochore

forms as a protein plaque on the surface of the centromere during late prophase. Spindle microtubules attach to the kinetochore on each chromatid

WHat occurs in early prophase

chromomes start to condense and the mitotic spindle begins to form and the nucleuolus disappears

What occurs with cohesin in the cell cycle

During s phase, the protein cohesin is assembled along the lengths of the chromatids, Polo-like-kinase 1 (Plk1) phosphorylates SA2 (subunit of cohesion ring), causing cohesion rings to disassemble everywhere except at the centromeres. By metaphase, cohesin is present just at centromere so chromatids are attached only at centromeres. Cohesin holds the dupplicated chromatids together until their separation anaphase

How does condensin help condense chromosomes in prophase

In late G2, M-Cdk phosphorylates condensin, which assembles on the DNA and helps condense it. The condensin forms a structural framework in the mitotic chromosomes and helps coil the DNA of the 2 chromatids

What does centrosome contain in G1

pair of centrioles

What occurs to centrosome and centrioles in S phase

dupicliate, but both parts of daughter chromosomes remain together until mitosis

What happens in early mitosis tow the centrosomes

the two centrosomes nucleate microtubules which push them apart - this is the beginning of the spindle. Centrosomes are connected by interpolar microtubules which grow out from each centrosome and overlap where they meet

What do centrosomes do as the nuclear lamina and envelope are breaking

the centrosomes move apart over the surface of the prophase nucleus. They form the poles of the spindle

What occurs to microtubules as cells transition from G2 into prophase

microtubules are reconfigured

Spindle microtubules versus cytoplasmic tubules and why

spindle microtubules are shorter and less stable because M-Cdk phosphorylates microtubules-associated proteins (MAPs)

what do less stable microtubules do

they readily assemble and disassemble at centrosomes, which is essential for forming the mitotic spindle and for mitosis to occur

How do you go from long “stable” microtubules interphase structure to a bipolar structure in mitosis

upon nuclear envelope breakdown, MT dynamics change dramatically, catastrophe increases and rescue decreases. IT USES NUCLEAR TRANSPORT MACHINERY

How does nuclear transport machinery help assembly of mitotic spindle

importin-beta inhibits microtubule assembly factors, RanGTP binds to importin-beta, microtubule assembly factors are released

What three types of microtubules make up the mitotic spindle

astral, kinetochore, and interpolar microtubules

What occurs in prometaphase

nuclear envelope disperses and plus ends of microtubules probe into the condensing chromosomes.

A microtubule initially attaches laterally to a kinetochore, the chromosomes slides along the microtubule to the pole then attaches end-on. The kinetochore on the chromosome is then captured by a microtubule from the opposite pole → this bi orientation is stable

What are microtubule-associated proteins

compilation of motor proteins and nucleation factors (cross-link microtubules)

How do prometaphase chromosmes move

they move constantly, especially when they are attached to just one pole. Even when both kinetochores are properly attached (bi-orientation) to opposite poles, a prometaphase chromosme will still oscillate, moving towards one pole and then to the other

What happens in late metaphase

chromosomes line up at the equator, half way between the poles

What occurs in the transition from metaphase to anaphase

At metaphase, the protein cohension holds daughter chromtids together at their centromeres. Metaphase transitions to anaphase when all the hcromosames are in place on the equator with all the kinetochores proerply oriented and attached. Then anaphase promoting complex destroys cohesin and chromatids seaprate

The anaphase promoting complex

tags an inhibitory protein (securin) with ubiquitin causing securin to be broekn down in a proteasome. Removal of the inhibitor activates a proteolytic enzyme (separase) which breaks down the cohesin. Cohesin is only present at the centromere after prophase. The spindle then pulls apart the chromatids and the chromatids separate (now they are each chromosomes)

What occurs in Anaphase A

kinetochore microtubules depolymerize at both ends. This shortening pulls the chromosomes to the poles

What occurs in Anaphase B

bipolar kinesin causes sliding between non--kinetochore microtubules which pushes apart the poles. Bipolar kinesin is anchored by its ATPas heads to two non-kinetochore microtubules originating from opposite poles. The heads move along the mirtoubules to the plus ends. Tubulin heterodimers are then added to the plus ends of the non-kinetochore microtubules to maintain the overlap between them as they push apart the poles. Also aided by the forces exerted by outward-pointing astral microtubules

What do astral microtubules do

they contact the plasma membrane via a dynein motor molecule and since dynein is a minus-end directed motor, it will pull the minus end of the astral microtubule with its attached centrosome towards the plasma membrane. The astral microtubules then shorten by losing tubulin heterodimers at plus end

What occurs in telophase

APC destroys M-cyclin, and in its absence, M-Cdk loses its acitivity and dephosphorylation occurs. This causes the nuclear lamina, envelope, and pores to reassemble in telophase, and many microtubules disassemble. Dephosphorylation also causes chromosomes to decondense in telophase so chromosomes resume transcription

What are the two trasnient cytoskeletal structures that mediate M phase in animal cells and what do they do

The mitotic spindle (microtubules) assemble first to separate the replicated chromosomes. The contractile ring (actin/myosin) assemble to devide the cell in two.

What occurs in cytokinesis

proteins are recruited to non-kinetochore microtubules that activate RhoA. RhoA drives formation of contractile actin filaments. Actin and myosin-II comprise the contractile ring which is assembled immediately below the plasma membrane.

What does contractile ring action do

forms cleavage furrow

What is apoptosis

programmed cell death

How do cell division and apoptosis balance tissue homeostasis

balanced: have healthy tissue

too much division, no apoptosis: larger size

too much apoptosis, no division: smaller size

Why is regulation of programmed cell death important

it’s needed for many development processes

hallmarks of apoptosis

bleb formation, collapse of cytoskeleton, fragmentation of DNA, disassembly of nuclear envelope, and cell surface alterations that attract macrophages for phagocytosis

How is apoptosis executed

intracellular cascade of caspase-dependent proteolysis:

Inactive initiator caspase has two inactive monomers both with adaptor-bidning domains, protease domain, and cleavage sites. An apoptotic stimulus of adaptor proteins leads to dimerization, activation, and cleavage. So, the active initiaor caspase activates by cleavage, the inactive executioner caspase. This activate executioner caspase then cleaves multiple substrates → apoptosis

How does activation of Bax proteins (Bcl2 family members) promote apoptosis

activation of Bax protein by DNA damage, releases cytochrome c from the mitochondrial intermembrane space through activated Bax/Bak which forms pores on the outer MT membrane. Cytosolic cytochrome c binds and assembles the adapter protein complex, the apoptosome, which recruitts procaspase-9. Lastly, the caspase cascade is triggered by activation of iniator caspase in the apoptosome.

What are the 3 extracelullar signals regulating (1) cell sruvival, (2) division, and (3) growth

(1) survival factors: promote cell survival

(2) mitogens: stimulate cell division (increase in cell #)

(3) growth factors: stimulate cell growth (increase in cell size)

What do survival factors do

act by binding to cell-surface receptors. The activate receptor activates a trqasncirption regulator in teh cytosol. This protein moves to the nucleus where it activates the gene encoding Bcl2, a protein that inhibits apoptosis

What do growth factors do

Extracellular grwoth factors increase the synthesis and decrease the degradation of macromolecules. This leads to net icnrease in macromolecules and therefore cell growth. Some extracellular signal proteins can act as both grwoth factors and mitogens, stimulating both cell growth and progression thru cell cycle, which helps ensure that cells maintain their appropriate size as they proliferate.

What do mitogens do to stimulate cell proliferation

they inhibit Rb protein:

Mitogens bind to the cell-surface receptors and activate intracellular singaling pathways that lead to formation/activation of G1-Cdk and G1/S-Cdk complex. They phosphorylate thereby inactivate Rb protein. The trasncription regulators are now free to activate transcription of their target genes, leading to cell proliferation

What is mytostatin and whtat does it do

an extracellular signal that inhibits muscle cells to increase in number and size

What happens in mytostatin KO

muscles cells grow in excess because both size and number of muscle cells increased

How does mitotic spindle form in prophase

2 centrosomes form the poles and organize the microtubules. The minus ends of the microtubules are adjacent to the centrosomes and the microtubules are dynamically unstable (growing and shirking). at the zone of overlap, interpolar microtubules from opposite poles interact via a bipolar kinesin molecule that connects interpolar microtubules and moves to their plus ends. Kinesin slides the microtubules past each other - pushing apart the poles. Addition of tubulin to the +.ends of the interpolar microtubules maintains the overlap

What does cohesin do

holds chromatids together at centromeres

How many microtubules attach to each kinetochore in mammals

20-40 microtubules