CH 14: Cellular Reproduction

Question 1

Two major phases of the cell cycle

Answer 1

mitotic (M) and interphase

M phase is a very small portion of the cell cycle

cytokinesis is not a part of mitosis, but follows after mitosis

preceding mitosis is G2 phase

G1, S, G2, M

Question 2

Catagories of cells

Answer 2

specialized cells that have lost their ability to divide
=> muscle cells, red blood cells

Cells that can be induced to divide
=> liver cells, lymphocytes

Mitotically active cells
=> epithelial cells, hematopoietic stem cells

Question 3

Cell fusion experiments

Answer 3

fgusion of G1 cells with S-phase cells resulted in G1 nucleus stimulated to undergo replication

fusion of G2 cells with S-phase cells resulted in G2 nuclei not stimulated to undergo replicaiton

implicaitons =MCM protein are exported out the nucleus after replication. After replicaiton they are not present, thus not in the G2 cells and cant undergo replication

Question 4

Mitotic factors induce chromosomal complation

Answer 4

M-phase HeLa cell fused with G1 PtK2 cells
resulted in G1 chromosome compaction. Note, M phase chromosome is HIGHLEY compacted, and G1 is not at all.
=> something in the mitotic cell to tell G1 cells to compact

fusion of Mphase Hela cell fused with Sphase PtK2 cell
=> S is very uncompacted, leads to S chromosome fragmenttation

fusion of M with G2 results in G2 compaction

Question 5

the role of Protein kinases for cells entering M phase

Answer 5

entry into M phase initiated by protein kinases

maturation promoting factor (MPF)

MPF activting is coorelated to cyclin, that is it is DEPENDENT
=> cyclin dependent kinases

Question 6

Discovery of MPF

Answer 6

early experiment with egg development= maturation process

progesterone stimulates first meiotic division from the primary oocyte to the ovum

“Germinal vesicle breakdown”

ratio of induced maturation to volume of injected cytoplasm is highest at 20 hours following progesteron treatment

took cytoplasm out of a fertilized frog egg (treated with projesterone) and put it into an unfertilized frog egg

Question 7

MPF activity cycles in amphibian eggs

Answer 7

MPF activity present in cytoplasm 90 minutes after fertilization

activity correlates with cell divisions (Cleavages)

Question 8

Cell cycle extracts from HeLa cells

Answer 8

germianl vesicle breakdown (GVBD) mazimal wtih mitotic extract

activity not present in extracts from early G1, late G1 or s phase

Mitotic extract from a somatic cell can influence egg cells to divide

thus, all cells have the same maturation promoting factor needed for that cell to ;divide

Question 9

Protien A vs protien B iin cell division

Answer 9

protein A = cyclin. cycles up and down by increaseing right before cell division then dropping off

Protien B keeps increasing over time

Question 10

MPF activity

Answer 10

purified MPF stimulated the incorporation of p32 into protiens and is thus a kinase

MPF consists of two polypeptides
32 kda and 45 kda
45 kda protein becomes labeled with p31 (cyclin)

Yeast produce 34 kda protein with kinase activity (cdc2)
=> antibodies from cdc2 react with amphbian 32 kda protein
=> highly conserved protein, same regualtion in all eukaryotic cells

Question 11

Cyclin A mRNA induces maturation

Answer 11

xenopus oocytes either injected with cyclin mRNA or stimulated with cyclin mRNA or stimuated with progesterone

percentage of oocyted exhibiting germinal vesical breakdown (GVBD) measured

Question 12

Cyclin dependent kinases

Answer 12

cdks first studied in yeast

temperature sensitive mutants idenify two stop points
=end of g1
=end of g2

fission yeast
=cdc2 responsible

Question 13

cell cycle regulation in yeast

Answer 13

cdc2/ g1/s cyclin phosphorylated ORC

cdc2/ m cyclin phosphorylated lamin proteins

CYCLINS ARE ARE SAME but the kinases are different

Question 14

regualtion of cdks

Answer 14

cyclin concentration
= differential transcription of cyclins at stages of cell cycle

controlled proteolysis
=cyclin concentration regulated by controlled synthesis link cyclins to polyubiquitin chain (degradation signal)

cdk phosphorylation state
=kinases CAK and Wee1
=phosphatase cdc25

Question 15

cdk phosphorylation state

Answer 15

CAK phosphorylated Thr161

Wee1 phophorylated Tyr15
=> inactivated cdc2 kinase

cdc25 phosphatase removes Tyr15 phosphate
=> activates cdc2 kinase

Question 16

yeast cell cycle mutants

Answer 16

wee1 mutant (kinase)= cell divides prematurely

cdc25 mutant (phosphatase)= cell continues to grow, doesn’t divide

Question 17

Subcellular localization of cyclin

[cyclin b1]

Answer 17

G2 to prophase [m]

G2, cyclin is in cytoplasm, is then imported into the nucleus for prophase
=> phosphorylation of nuclear expport signal (NES) stops export form nucleus

Question 18

Mammalian cyclins and cdks

Answer 18

Cyclin b/a + cdk1 initiates G2 to M phase

cyclin D’s + cdk4 cdk6 activates pRB which increases transcription of cyclins E and A

cyclin E + cdk2 initiates G1 to S phase

Cyclin A + cdk2 initates S to G2

___________________________________

cdk1 is absolutelyy required for development

mice expressing only cdk1 and cdk4 develop into sterile adults
=> defects in meiosis

Question 19

Cell cycle checkpoints

Answer 19

delay profression of cell cycle if
=>chromosomal DNA damaged
=> DNA replicaiton not completed
=> misalignment of chromosomes

requires three classes of proteins
=> sensors that detect abnormaliates and emit signals (often kinases
=> Transmitters that send signal along pathway
=> effectors that respond to signal and inhibits cell cycle machinery

Question 20

DNA damage checkpoints

Answer 20

G2 check
=> ATR sensor is activated by ultraciolet damage (thymadine damage) activates chk1 and then phosphrylated cdc25. cdc25 when phorphorylated in inactive

G1 check
=> ATM sensor is activatd by ionizing radiation (double stranded breaks) and activates chk2 which phosphorylated p53 and activates p21 transcription
=> p21 protein directly inhibits cdk

Question 21

cdk inhibitor arrests cell cycle progression

Answer 21

p27 (similar to p21) alters conformation of cdk catalytic subunit

=> inhibits protein kinase activity

Question 22

gentics of cancer [chapter 16]

Answer 22

cancers are often monoclonal
=> arise from a single cell

Malignancies require more than a single genetic alteration

inheritance of mutated genes is not the leading cause of cancer (shown in twin studies= envriomental hazzards)

Stem cells may be the source of tumors
=> natural selection for cells that have advantanges
=> i.e., telomere length

Question 23

Genetic alterations for cancer

Answer 23

expression of telomerase

epigenetic changes
=> promoter alteration
=> methylation patterns

mutations of tumour suppressor genes
=> p53

oncogenic genes
=> growth factor receptors, and associated genes

Question 24

tumor cell types depend on what type of tissue they arise from

Answer 24

x = tissue stem cell

y = pluripotent progenitor cell

z = commited progenitor cell

tumor cells can arise from stem cells or progenitor cells

Question 25

Precancerous cells often exhibit histological changes

Answer 25

normal cervial epithelial cells have small nuclei and homogenous in shape abnormal cells from cervixal carcinoma with large nuclei, heterogenous in shape (think of wee1 mutant, quick division)

Question 26

Tumor suppressor gene // and // oncogene

Answer 26

both copies of tumour suppressor gene must be knocked out mutationin one of two copies of oncogene sufficient (dominant)

Question 27

Types of mutations that lead to cancer cel cycle

Answer 27

altered protein strucutre geneamplification - overexpression rearrangement of DNA alters expression of strucuture figure 16-12

Question 28

Mutations in Rb gene lead to Cancer

Answer 28

Sporadic = individual begins with two normal alleles Familial = individual inherits one mutated gene

Question 29

Role of pRb in cell cycle

Answer 29

pRb bound to E2F protein => Trancription repressor for genes involved in G1 to S progression Cdk phosphorylated pRb => Dissocated from E2F E2F activates transcription

Question 30

Role of p53 in cancer and the cell cycle

Answer 30

Tumor suppressor gene => p53 levels rise in response to DNA damage 5o % of human cacers linked to mutation in p53 gene Activates p21 gene whose product inhibits progression form G1 to S => Cell repairs damage p53 can promote cell to enter apoptocitc pathway => activates Bax gene

Question 31

p53 mutations

Answer 31

frequency of mutations for p53 aminos acids removal of positively charges Arginine whihc prevents the p53 TF from binding to the DNA backbone most frequents mutations correleated with regions interacting with DNA

Question 32

Role of p53

Answer 32

p53 expression increases in response to DNA damage => arrests cell in G1 or directs cell to the apoptotic pathway p53 mutatuion consequences => mitotic failute => Division with damage => tumour ``` double p53 (-/-) mutants less susceptilble to chemical// chemothreaputic treatment. => continue to divide and do not become apoptotic ```

Question 33

p53 and senescence

Answer 33

Senescence => cells remains alive and metabolically active (non-dividing) Senescence can be triggered by Ras oncogene (ex benign mole) Inactivaiton of p53 leads to malignalcy => tumour progression

Question 34

p53 and PST

Answer 34

posttranslational modificaitons of p53 regulate the specificity of the enzyme phosphorylation, acethylaiton, methylation alternateive splice variants

Question 35

sensecence and tumorigenesis

Answer 35

cellular senescence triggered by one of two major tumour suppressor pathways, p53 and pRB cells are blocked at the G1/ S transition Senescent cells accumulate in aged tissues

Question 36

Senescene associated secratory proteins (SASP)

Answer 36

have multiple effects = inflammaiotn = enchance proliferation of tumours Platelet cells and tumorus interact = tumours activate platelets =activated platelets release factors that affect proliferation or metastasis of tumours interactiosn of senescent cells and platelets? = alter the functional status of platelets and fibrinolysis

Question 37

Familial adenomatous polyposis coli (FAP)

Answer 37

inherited disorder that leads to polup formation on colon deletion of APC gene on chromosom 5 => one allele mutated - premalignant => two alleles mutated - malignant APC = anapahse promoting complex loss of APC functinon leads to abnormal chromosome segrefations = aneuploydy APC gene muations = premaligants polyps in colon epithelial cells

Question 38

Breast cancer

Answer 38

BRCA1 and BRCA2 genes linked to high proportion of breast cancers => respond to DNA famage and initiate repair

Question 39

Proto-oncogenes

Answer 39

oncogenes promote loss of growth control and conversion to malignant state approximately 100 protooncogens have been identified => mostly linked to processes involved in cell growth => often included in genomes of RNA tumour viruses ex: sis viral oncogen derived from PDGF : erbB encodes EGF receptor

Question 40

Genomic landscape of colorectal cancer

Answer 40

APC and TP53 are common mutations there are also many different unique somatic mutations 70% of colon cancers exhibit chromosome instability and aneuploidy

Question 41

mitosis

Answer 41

process of nuclear division that faithfully partitions duplicated cvhromososmes five stages prophase, prometaphase, metaphase, anapahse, telophase

Question 42

cytokinesis

Answer 42

divison of cell that partitions cytoplasm into two cellualar packages

Question 43

Prophase

Answer 43

chromosomal material condenses to form compact chromosomnes cytockeleleton is disassembled and mitotic spindle is assembled golgi complex and ER fragment. Nuclear envelope disperses (triggerd by phos of lamin proteins)

Question 44

Prometaphase

Answer 44

chromosomal microtubesla attach to kinetochors of chromosomes chromosoems are moved to spindle equator

Question 45

metaphase

Answer 45

chromosomes are aligned along metaphase plate attached by chromosomal microtubules to both poles

Question 46

anaphase

Answer 46

centromeres split, and chromatids separate chromosomes move to opposite spindle poles spindle poles move farther apart

Question 47

telophase

Answer 47

chromosomes cluster at opposite spindle poles chromosoems become dispersed nuclear envelope addembles aroung chromosome clusters golgi complex and ER reforms Daughter cells formed by cytokinesis

Question 48

Condensin and Cohesin

Answer 48

condensin forms loop around DNA loops Cohesin forms loop around sister chromatids => alternative model: cohesin forms moleucular bridges between sister chromatids

Question 49

Mitotic chromosomes

Answer 49

cohesin dissociated from sister chromatid arms during prophase => polo-like kinase and aurora B kinase phophorylate cohesin remains at centromere

Question 50

Kinetochore structure roles

Answer 50

roles: site of attachment to mitotic spindle location for motor proteins components fo signaling pathway for cell cycle checkpoint

Question 51

Kinetochore structure proteins

Answer 51

Dynein moves toward minus end of microtubule CENP-E (kinesin) moves toward plus end of microtubule NDC80 is a multisubunit protein that connects microtubule to kinetochore depolymerase = specialized kinesin protein responsible for depolymerizing mt during anapahse

Question 52

Cohesin

Answer 52

dissociated from sister chromatid arms during prohpase* remains at centromere polo-like kinases and aurora B kinase phophorylate cohesin phosphatase at centromere removes phosphates groups to maintain cohesin

Question 53

Roles of kinetochore structure

Answer 53

1 site of attachment to mitotic spidle 2 location for motor proteins 3 component of singaling pathway for cell cycle pathway

Question 54

kinetochore strucute the proteins

Answer 54

Dynene = moves toward minus end of microtublules CENP-E (kinesins) moves toward plus end of mt deplumerases-specilized kinesin protein responsible for depolymerizing mt during anaphase

Question 55

Formation of mitotic spindle centrosome cycle see refrence for picture

Answer 55

phosphorylation of centrosomal protein by cdk2 initiates duplication centroles duplicate in s phase centrosomes migrate to opposite poles in mitotis G1 pericentrioloar nuceation site S duplication G2 elongationa nd maturation M centrosome seperation (Holland et al 2010)

Question 56

formation of mitotic spindle prophase

Answer 56

during prophase-spindle microtubules start to organize and emenate from centrosomes microtubules = astral microtubules => these go out to the end of the cell and play an impirtant role in cytokeneisis = form the cleavage furrow

Question 57

Abnormal centrosome number

Answer 57

leads to abnormal chromosome segregation

Question 58

spindle formation in absence of centrosomes (as in plants) ***

Answer 58

spindle fibres nucleated at chromosomes instead of centrosome minus end directed motor proteins have multiple heads that cause minus ends of chromosomes to converge multiple heads on one protein linkes and pulls microtubles together to make a spindle pole

Question 59

early prometaphase

Answer 59

immediately following nuclear envalope breakdown chromosome become associated with plus end of microtubules chromosomes associated randomly on a microtubule, but the motor protein activity of the motor-protein arm will direct the chromosome towards the tip of the microtubule where it will become bound

Question 60

tension

Answer 60

when a chromosome is equally lined up on the prometaphase plate, there is equal tension on either kinetochore important signalling process equal tension on all chromosomes means that the cell can progress towards anaphase

Question 61

role of motor proteins

Answer 61

congression of chromosomes to the metaphase plate deficient for kinesin protein associated with the + directed chromosoems samres chromosomes can not congress motor protein normally provides force for moving away from arms

Question 62

microtubule behaviour

Answer 62

differential polymerization rattes regulated by differntial pulling force (tension) on chromosomes chromosome tethered by motors (and ndc80) tension = treadmilling => equal rates of polymerization and depolymerization (figure 14-23)

Question 63

Mitotic spindle of animal cells

Answer 63

minus ends of all spindle microtubules oriented towards centrosomes anstral microtubules = help position the spingle apparatus, may help determine plane of cytokenesis chromosome microtubules = exert pulling force polar microtubules = maintain mechenical integrety of spindle

Question 64

Tubulin Flux at metaphase

Answer 64

treadmilling tubulin monomers incorperated at plus ends (kinetochores) (net gain) monomers lost at minus ends (centrosome) (net loss)

Question 65

anaphase control

Answer 65

APC and SCF responsible for adding ubiquitin and targeting protiens to proteasomes (APC = anaphase promoting complex) APC^Cdc20 labels securin protein (metaphase/ anaphase) APC^Cdh1 ubiquinates mitotic cyclins (pregresses to g1) Cdh1 activity is inhibited by Cdk1 mediated phophorylation during mitosis. Becomes active as cdk1 activity drops Destruction of mitotic cyclins promotes mitotis to G1 transition Destruction of securin actvates "separase" enzyme that cleaves cohesin protiens- sister chromatids separate

Question 66

Importance of proteolysis in mitotis

Answer 66

cdk1 inhibitor added mitotis complete, initiate cytokinesis cdk1 is progression into mitosis. washed free of cdk1 inhibitor, cdk1 is now active and cell goes back into mitosis becuase cyclin b was not degrated. cell thus enters mitosis again need to degrade proteins like cyclin B to regulate teh cell cycle

Question 67

events of anaphase

Answer 67

anaphase A => monomers lost from plus ends of chromosomal microtubules => seperation of sister chromatids anaphase B => monomers added to plus ends of polar microtubules => pushes poles apart => poles move apart and sister chromatids (now chromosomes) move to the poles) polar microtubules become larger, push past each other and moves the poles furtherapart kinetochore attachement we see depolymerization which pulls chromosomes to the opposite ends

Question 68

micotubule depolymerization

Answer 68

catastrophic dissasembly of the microtubules causes chromosomes to move toward the basal body= organizing center basal body-microtubule organizing center as tubulin dimers are diluted fromsolution, microtubule ahortens and pulls chromosome toward basal body

Question 69

catastrophic disassembly of microtubules

Answer 69

chapter 9-25 GTP dimers are incorperated into tubulin GTP to GDP dimers in you increase the rate of hydrolysis or lower teh rate of GTP dimers, the rate of hydrolysis catches up to the rate of incorperation// polyermerisation the flared out strucutre is more stable that linear, flared out GTP dimer structure is overtaken => catastrophic dissasembly

Question 70

Microtubule depolymerization (anaphase)

Answer 70

depolyermization of microtubule at both ends depolyermizing kinesins at either end of microtubules

Question 71

Spindle (assembly) checkpoint

Answer 71

chromosome fails to attach to spindle fiber attaches and moves polewards until stable spindle checkpoint protein - Mad2 labeled pink =only found on kinetochores that are mod attached to spingle fiber = mad2 binds cdc20 not able to ubiquinate securin mad2 blocks cdc20// degredtion process => no seperation

Question 72

Importance of mechanical tension

Answer 72

aurora B kinase enzyme (activity at centromere) responds to lack of tension phophorylates unknonw substrate destabilizes MT attachment kinetochores reattach

Question 73

Activity of motor protein (prometaphase)

Answer 73

1 plus end directed motor proteins cause polar mictobuluels to slide past each other 2 minus end directed motor protein at kinetochore 3 plus end directed motor protein associated with kinetochore and chromosomes

Question 74

Activity of motor protein (metaphase)

Answer 74

4 plus end directed motor activity on polar microtubules 5 balanced activity of motor proteins at kinetochore

Question 75

Activity of motor protein (anaphase)

Answer 75

6 kinetochore motors move chromosomes and/or anchor chromosomes to depolymerizing microtubules 7 motor proteins separate poles

Question 76

cytokinesis

Answer 76

actin micofilaments at equator of dividing cell | => myosin is important too

Question 77

role of myosin in cytokinesis

Answer 77

starfish egg incubated with myosin antibody whihc inhibits cytokinesis mitotis continues in absence of cytokinesis the two processes can be seperated

Question 78

position of spindle pole

Answer 78

cleavage plane forms between spindle poles narrow cell divides quicker cleavage furrow is at the midpoint between the centrosomes => they are responsible for formaiton// alignment of the cleavage furrow

Question 79

Position of spindle pole

Answer 79

DIntance from poles to cleavage plane is smaller in cylindrical cell shortens time it takes the cleavage signal to reach the surface (cortexz) equatorial stimulation

Question 80

models to explain how the mitotic apparatus signals the cortex to determine the poistion of the cleavage furrow

Answer 80

Astral relaxation => signals from teh astral centers induce relaxation of the cortex near the poles Equatorial stimulation => by signals from the astral centers. Signals from the astral centers induce contraction of the equatorial cortex Equatorial stimulation => by signals from the spindle midzone. Signals from teh spingle midzone induce contraction of the equatorial cortex

Question 81

Cytokinesis in Plant cells

Answer 81

organization of microtubule arrays changes during M phase preprophase band-site of cleavage phargmoplast-role in formation of cell wall\ 1 folgi derived vesicles organized on cell plate 2 fingers extended that fuse with neighbouring vesicles 3 membranes fuse with original plasma membrane contents of vesicles form cell plate

Question 82

meiosis

Answer 82

mechanism to reduce chromosome number by half two sequential cell division without an intervening rounds of DNA replication division 1: homologous chromosoems separated division 2: sister chromatids divide into separate chromosomes

Question 83

prophase 1 of meiosis`

Answer 83

pairinng of homologous chromosomes

Question 84

anaphase one of meiosis

Answer 84

metaphase 1 lining up of chromasomes anaphase 1 separation of homologous chromosomes

Question 85

telophase 1 of meiosis

Answer 85

homologous chromosomes migrate to opposite poles`

Question 86

prophase 2 of meiosis

Answer 86

second meiotic division identical to mitosis

Question 87

gametic or terminal meiosis

Answer 87

all multicellular organisms primary role is formation of gamates

Question 88

zygotic or initial meiosis

Answer 88

protists and fungi meiotic divisions occur just after fertilizatiohn to produce haploid spores spores mitotically divide to produce haploid adult generation

Question 89

sporic or intermediate meiosis

Answer 89

plants and some algae meiosis not used to produce gametes gametes produced by mitotic divisions of haploid gametophyte

Question 90

see figure

Answer 90

14-20

Question 91

meiosis and alternate life cycles

Answer 91

gametic and sporic = fusion of egg and sperm, diploid adult zygotic = meiosis immediately after fusion of haploid spores sporic = sporophyte meiotically divides to produce haploid spores zytoic = spores divide mitotically to produce haploid adult generation sporic = gametes formed from gametophyte by mitotic division zygotic = haploid spores generated from mitotic division of adult

Question 92

meiosis in vertebrate males

Answer 92

meiosis occurs before differentiation of sperm produces four viable gametes spermatogonia => spermatocytes (primary) => secondery => spermatids spermatids differnitate to become sperm

Question 93

meiosis in vertebrate females

Answer 93

meiosis occus after differentiation of the primary oocyte produces only one fertile egg primary oocyte arrested in prophase I until ovulation

Question 94

stages of prophase I

Answer 94

leptotene = condensed chromosomes first visible zygotene = visivble associateion of homologues (synapsis) pachytene = end of synapsis diplotene = dissolution of synaptnemal complex, chiasmata visible diakinesis = meiotic spindle visible

Question 95

end of leptotene

Answer 95

telomers organized together in a bouquet and associated with nuclear envelope

Question 96

synaptonemal complex

Answer 96

complex that holds the homologous chromosomes togeter lateral and transverse complexs

Question 97

when does crossing over occur

Answer 97

occurs during beginning of diplotene chiasmata are the crossing over structures

Question 98

separation of homolgous chromosomes and sister chromatids

Answer 98

sister chromatids held together by cohesins [synaptonemal complex] homologues held together by chiasmata because after the synaptonemal complex breaking down entering metaphase I cohesin cleaved between arms remains at centromeres chromosomes seperate metaphase II cohesisn breaks down and sister chromatids separate oocyte arrested at metaphase II due to inhibitions of APC^cdc20 fertilization leaads to calcium influx and activaiton of APC^cdc20 and destruction of cyclin B

Question 99

meiotic nondisjunction

Answer 99

20-25% of human oocytes are aneuploid (monosomic or trisomic) primary nondisjuction = incorrect segregation of homologous chromosomes (two gamates with extra, two with none) secondary nondisjunction = incorrect segregationo f sister chromatids (two normal gamates, one with additional, one with nothing) more prevelant in older egg cell because the chiasmata can let loose early and lead to early seperation of homologues chromosome 21 trisome survies unlike many other due to gene dosage= smaller chromosome

Question 100

turner syndrome

Answer 100

XO female with genital development arrested and ovaries fail to develop

Question 101

klinefelter syndrome

Answer 101

XXY male with presence of feminine physical characteristics

Question 102

XYY

Answer 102

physically normal male, taller than average, supermale

Question 103

genetic recombnination

Answer 103

echange of maternal and paternal alleles during meiotic prophase I physical breakage of DNA molecules and religation with split ends of homologous chromosomes involves DNA repair enzymes that fill gaps furing exhange process

Question 104

genetic recombnination process

Answer 104

double strand bvreak in one chromosome widening of gap by exonuclease activity (5' to 3' excision) single strand invades homologous chromosome (Rad51 protein) and displaces strand DNA repair using the homologous strand as a template heteroduplex formation with a holiday junction breakage and lreligation with resolvase enzyme to make with cross over or non-crossover products see nexus video