Week 3 Chromatin/Epigenetics Flashcards
what structural motif allows specific histone interactions
- Globular domain contains a histone fold motif that provides the basis for dimerization interface– handshake interaction
describe structure/components of nucleosome
smallest repeating unit of chromatin
- 147 bp DNA wrapped around an octamer of histones
4 distinct histones– 2 of each type
briefly explain the use of MNase sequencing in mapping chromatin structure
what is the end goal (think occupancy)
- exo/endonucleases used to digest naked DNA betw/ Nucleosomes
– remaining undigested DNA subjected to high throughput seq.— mapped on reference
gives idea of nucleosome occupancy on DNA
in what do transcriptional activators (which bind to enhancer cis-regulators sequences) facilitate the altering of local chromatin structure?
- T activators attract coactivators ATP-dep chromatin remodellers, histone-mod enzymes, histone chaperones
how do transcriptional activators bound to enhancers 1000s of KB away from promoter facilitate assembly?
- bonus: what is the role of mediator complex in this?
DNA looping– brings T activator into close proximity to promoter
— mediated by MEDIATOR protein—ties interaction between T activators, RNAP, GTFs and their assembly at promoter
how does histone chaperone mediated replacement and removal of histone affect DNA access?
REMOVAL– allows assembly of transcription machinery on histone free DNA
REPLACEMENT —shifts chromatin structure so there is greater access to nucleosome DNA
How does DNA methylation repress transcription/ promote heterochromatin formation
- methylated DNA acts as binding site for histone modifying enzymes– induce heterochromatin conformation
Xist (along with Tsix) is produced from XIC cis-acting region of X chromosome.
give examples of 1ry/2ry silencing factors induced by Xist and how they induce changes in chromatin structure (i.e. lead to X chromosome compaction)
1ry
Depletion of acetylation, H3K4me (at enhancer site)
- accumulation of H3K27me (facultative heterochromatin)
2ry
DNA methylation of genes promoters– H3k4me3
when are chromatin structures disassembled in cell cycle
S phase
what enzyme facilitates maintenance methylation i.e. methylation of hemi-methylated DNA strands (right after daughter strand synthesis)
- what accessory protein needed in vivo?
what process is DNA methylation coupled to (hint: H3K9me3)?
- maintenance methylation facilitated by DNMT1— preference for hemi-methylated DNA where one strand has unmethylated CpG site
- maintenance methylation in vivo requires UHRF1 protein
coupled to Histone methylation via Suv39 writer which methylates K9 on H3
outline the positive feedback mechanism between HP1 (reader) and Suv39H1 and the outcome
HP1 reader recruits Suv39 and facilitates methylation of neighboring chromatin at H3K9 sites– induces repressive chromatin form
– way by which heterochromatin is established
distinguish between direct histone modifications acetylation and methylation
acetylation– reduces histone positive charge—disrupts electrostat. Histone-DNA interaction– less compact chromatin—DNA ACCESS
methylation– promotes positive histone charge– leads to more compact chromatin
describe the result of histone chaperone-mediated replacement of H2A histone with H2A.Z on chromatin structure and overall gene transcription
- think of nucl. barrier, elongation
dimers including H2A.Z are more easily lost during nucleosome unwrapping (which occurs ahead of repl. fork) — LOWER NUCLEOSOME BARRIER
—that DNA becomes accessible— RNAPII elongation
how do the following chromatin remodeling complexes affect chromatin access?
- SWI/SNF
- ISWI/CHD
- INO80
SWI/SNF- facilitates chromatin repositioning– nucleosome ejection/dimer eviction
INO80–facilitates histone exchange
ISWI/CHD– nucl. maturation/spacing
what modifications do the following domains recognize?
- Bromo domain
- Sant domain
- chromo domain
- acetylation on K
- phosphorylation of K, T residues
- methylation of K
specific PTM histone modifications can be found more often in specific regions
- where are the following mods found?
- H3K4Me3
- H3K27Me3
- H3K9Me3
- at promoter
- facultative heterochromatin
- repeats– constitutive heterochromatin
what common features of chromatin remodelers allows them to physically move DNA
- ATPase motor that breaks DNA histone contacts (H bonds)
name of the domain of HP1 that allows it to read H3K9me3/ IN POLYCOMB H3K27me3
chromodomain
briefly describe Polycomb repressive complex and the function of PRC2 protein in methylating H3
- repressive complex present in higher eukaryotes
- PRC2 component conducts H3K27Me3 on new daughter strands using the H3K27 of old histone
affect of DNMT1 on SUv39 (and hence histone methylation) at replication fork?
DNMT1 recruits Suv39 so that DNA AND histone methylation are coupled (positive feedback loop)
result of nucleosome disruption ahead of replication fork?
- effect on H3-H4?
- what essential chaperone (also implicated in transcription) mediates histone disassembly/re-assembly behind fork
FACT chaperone mediated H3-H4 tetramer–>dimer– loaded back onto daughter strands–
why is histone recycling important?
- think about the modifications on the histone and what they mean
modified histones contain important information— must be transmitted to daughter strands
outline reader and writer of PRC2 complex in propagating histone H3k27Me3 modification during nucleosome recycling
reader– EED
- writer–EZH2
what is a common mutation in pediatric cancer that causes defective silencing establishment?
– i.e. reader, writer is blocked/ not permitted to move forward
– H3K27–>H3M27
risk of epigenetic cancer therapy i.e. use of DNMTi, HDACi?
(note: DNMTi,HDACi are already accepted for clinical use)
while trying to reverse silencing of tumor repressors in cancer
- you do not control what is re-expressed – could be dangerous
