oncol mt 1 pt 2

tumor suppresor genes role

inhibit cell proliferation and survival

what happens when TSG is completely or partially lost

increase likelihood of cancer development

is activation of oncogenes adequate to cause cancer? if not, what else is involved

NOOOOO, TSG are also dysregulated in cancer

loss of TSG happens more frequently that activation of proto-oncogenes

retinoblastoma what and origin

tumor of retina from photoreceptor cells (stem cell precursor of cones)

sporadic retinoblastoma

non heritable

single eye tumor (unilateral)

after treatment, no futher risk of other tumors

familial form retinoblastoma

heritable bilateral tumor in retina

after treatment, increase risk of bone and other cancers

Rb

TSG gene involved in retinoblastoma

mutations are recessive and cause cancer

show with DNA the difference between sproadic and familial retinoblastoma

two mutations is much more rare than one

deletions in retinoblastoma

Rb is deleted

what happens when one TSG allele is mutated

it becomes inactive, and the other one compensates

if they both become mutated they transform cells

why are sequence mutations not likely and what is more plausible

one mutation is 1/10^6 two mutation would mean 1/10^12,,,,

1. translocations and deletions (meh)

2. epigenetics (gene silencing)

3. one copy mutated second copy altered to same mutation

examples of where one copy of a gene is mutated so the second one alters

mitotic recombination (more common) + gene conversion (most common)

mis-segregation at mitosis or deletion (less common)

mitotic recombination

2 homologous chromosomes exchange DNA during mitosis in somatic cells

LOH

loss of heterozygosity

when a heterozygous gene becomes homozygous dominant or recessive

gene conversion

homologous chromosomes

chromosomal mis-segregation

during mitosis

DNA methylation

add methyl group to cytosine at CpG site by DNA methyltransferase

heritable epigenetics

effect of methylated CpGs in DNA

1. recruit protein complexes that functions as histone deacetylase

2. removal of acetyl groups from histone in chromatin

3. more densley packed chromatin (inactive)

4. transcription stop

what are the 2 mechanisms that inactivate both copies in promoter methylatio

both copies are methylated independently (less common)

one copy is methlylated, second copy lost through duplication of methylated copy

2 types of methylation in cancer

global hypo: dec DNA methylation of highly repeated sequences = instability

localized hypermethylation gene silences

how are oncogenes identified

transformation assay

why are biological identification of TSG more complicated

TSG prevent transformation, so its harder

how can we ID TSG

genomics

ID region where LOH happen → find TSG here

SNP in TSG finding

SNP can help us distinguish alleles, distinguished can help us map sites

gatekeepers

½ proteins encoded by TSG

direct control by hindering cell proliferation (cell division)

increasing cell differentiation and/ or death

caretakers

½ protein encoded by TSG

maintain integrity of genome

two types of bad miRNA

1. no miRNA = proliferation of oncoprotein… bruh (loss of miRNA suppresor)

2. miRNA inhibits TSG from producing cancer killers …. bruh (oncogenic miRNA)

what do gatekeepr proteins respond to

growth inhibitory signals

DNA damage

unkown :o

familal adenomatous polyposis

heritable susceptibility to develop adenomatous polyps in colon → colon carcinoma

linked to loss of tumor suppressor APC

APC

tumor suppressor gene commonly found inactivated in sporadic colon carcinomas

why is APC an important tumor suppressor

wild type vs mutated intestinal crypt

what is the most important molecular consequence of APC inactivation

beta-catenin accumulation…

percentage of sporadic colon carinomas

90% = APC inactivation

10% = wild type APC is inactivated by promoter methylation

beta-catenin mutations that cannot be phosphorylated for GSK degredation

cell cycle clock what and where

what: network of interacting proteins

where: nucleus

cell cycle basic

prophase

chromosmes condense

centrosomes assemble

metaphas

align on metaphase plate

mitotic spindle

anaphase

chromatids pulled apart by mitotic spindles

telophase

chromosomes decondense, nuclear membrane forms, cytokinesis begins

general checkpoints of mitotsis

which period are cells responsive to mitogenic GF and TGF-beta

green and yellow arrow

green: where cells are responding to mitogenic GF and TGF-beta

yellow: R point

when does R point occur

late G1

several hours before G1/S transition

what happens at R point

→ divide (continue) or no (g0)?

CDK

cyclin dependent kinase (serine/threonin kinase)

always present

cyclins

regulate CDK → activation + substrate specificity

cyclin + CDK combo in each part of cell cycle

cyclin amount graph

cyclins are degraded within minutes

how is each cyclin-CDK activated and how is it shut down

activated: D + 4/6 = mitogens (only extracellular part), the rest by the subsequent phase

shuts down: by predecessor

how is cyclin D activated

extracellular signals: ECM (Fak), GF, Wnt, others

how are other cyclin families (EAB) activated

regulated by cell cycle

intracellularly

CDK inhibitor names and what do they inhibit

where must CDK inhibitors be to inhibit cyclin/CDK

nucleus

p15 activation and what stage

early and mid G1

p21 activation and what stage

almost every stage

how are CDK inhibitors inactivated

CDK can only be active in nucleus

explain this image

pRb origin and main goal

encoded by RB TSG

guardian of R-point gate

phosphorylation cycle of pRb

after mitosis, pRb is (hypophosphorylated/ dephosphorylated/ hyperphosphorylated) by _______

dephosphorylated; protein phosphatase 1

in early and mid G1, pRb is (hypophosphorylated/ dephosphorylated/ hyperphosphorylated) by _____

hypophosphorylated; cyclin D- CDK 4/6

after R point pRb is (hypophosphorylated/ dephosphorylated/ hyperphosphorylated) by

hyperphosphorylated by EAB CDK complexes

in S and G2 phase phosphorylation of pRb can be ______ by ______

transiently reversed by unknown phosphatases

diagram of pRb being hyperphosphorylated

pRb and E2F interaction

what is E2F and what are its target genes

E2F= transcriptional activators

targets: DNA precursor nucleotide synthesis, and DNA replication

positive feed back loop of cell cycle

3 ways pRb can be inactivated

1. mutation of RB gene followed by LOH

2. methylation of RB gene promoter = silencing

3. inactivation of pRb by HPV E7 proteins (human cervical caner)

how does inactivation of pRb by HPV E7 work

E7 proteins bind strongly to pRb, prevents E2Fs from binding, E2F constant activity

cyclin D1 _____ in cancer by

expression is elevated; Ras/Raf and PI3K and AKt signalling paths

(increased transcription and reduced degradation)

correlation between cyclin levels and deaths

myc reulgation of cell cycle

also promote degradation of CDK inhib (Miz)

5 molecular changes leading to deregulation of cell cycle clock

1. inactivation of pRb

2. overexpression of cyclin

3. increase activity of CDK (activation mutation)

4. improper localization of underexpression of CDK inhibitors

5. alterations induced by Myc

explain this picture

initiallt, p53 was thought to be an ongogene

however, cDNA had point mutation

most mutations carried by p53 gene are______ that lead to ______

missence point mutations; amino acid substitutions

dominant negative

p53

a mutant allele of a gene interferes with wild type when co-expressed (becomes dominant)

majority of p53 mutations affect where

DNA binding domain

not tetramization

in most mutations, how is wild type p53 discarded

through LOH

finish

half life of p53

20 minutes

signals that increase p53 protein level

radiation ______ p53 levels which _____ p21

increases; induces → cell cycle arrest at almost every stage

cytostatic function of p53

pro-apoptotic function of p53

p53-/- were resistant to irradiation induced apoptosis

Mdm2 mechanism

induce p53 ubiq. + degradation

inhibit p53 transcriptional activity

what happens when there is cell damage vs cell survival signals

where does Mdm2 and p53 complex go to get degraded

nucleus

high E2F induces what protein

p14ARF (human) and p19ARF (mice) binds to Mdm2 (TSG)

sequester in nucleus → cannot bind p53 → increase p53 → apoptosis

p 53 targets + mechanism

p53

TF and regulates gene expression

how is p53 anti-angiogenesis

induces expression of thrombosponding (TSP1) antiangiogenic protein

familial cancer symdrome

mother and father have one mutated copy of p53 (point mutation) → leads to very common family cancers

apoptosis plays important role in _____ that do not involve ______

physiological processes; p53

mitrochondrial path way

aka stress-activated pathway

apoptosis that is dependent on mitochondrial release of cytochrome C

death receptor pathway

apoptosis that does not require cytochrome c release (but it can accompant activation)

what is on mitochondria in relation to intracellular signals

balance between pro-survival and pro-apoptotic proteins on surface

what happens if apoptotic signals predominate on mitochondrial surface

1. channel in outer membrane opens

2. cytochrome c released in cytosol

3. activation of cascade proteases → caspases

4. cleavage of “death substrates”

5. apoptotic phenotype

pro-survival gene family

bax family

pro-apoptosis family #1

BH3 only family

pro-apoptosis family #2