Lecture 9: Chromatin and Gene Expression

Chromatin Remodelling

An ATP-dependent process altering DNA-histone interactions non-covalently that frees a binding site for a transcription factor

Chromatin Remodellers

Enzymes involved in chromatin remodelling

Chromatin Modification

Introduces covalent modifications on the histone tails

Swi/Snf

A chromatin remodelling complex that has a highly conserved ATPase subunit with 7 conserved motifs total — also contains a bromodomain

Swi/Snf, ISwi, CHD, and INO80

The four families of chromatin remodellers include:

Of the chromatin remodellers: ___ and ___ antagonize each other

ISWI

Chromatin remodeller that promotes regular nucleosome spacing (repressive)

Swi/Snf

Chromatin remodeller that creates open stretches of chromatin (activating)

H2A.X

H2A variant that has a serine 139 which is important for phosphorylation in the event of dsDNA breaks

repair proteins and cohesin

S139 phosphorylation helps recruit…

H2A.Z

H2A variant that is 60% identical to H2A and is involved in gene expression and chromosome segregation

SWR1

Enzyme that mediates the switch between H2A and H2A.Z

-1 and +1 nucleosomes, +1, -3 to +3, TATA-less promoters, open promoters, II, III, easily rejected from DNA, transcription

H2A and H2A.Z differ mainly in the ______— in yeast H2A.Z is found in the _____, in Drosophila _____, and in humans ______. H2A.Z is highly enriched in ______ and is preferably found in _____ (type ___ and ___). It is more ______ and promotes _____

10.5

The helical pitch is about ___bp per turn

translocase domain binds nucleosome positions 4-5, DNA from the linker region is pulled into the nucleosome and the displaced DNA rebinds position 1-5 forming a DNA loop over position 6, the DNA loop propagates away from the translocase domain, translocase resets and binds new DNA in position 4-5 and displaced DNA wave moves towards distal linker until distal linker is reached

Chromatin remodelling mechanism:

HAT

Histone acetyltransferase

HMT

Histone methyltransferase

HKMT

Histone lysine methyltransferase

HAT, HMT, HKMT, kinase

Writer proteins include:

Bromodomain

Possible domain in reader protein that binds acetylated lysines

Chromodomain

Possible domain in reader protein that binds methylated lysines

TUDOR

Possible domain in reader protein that binds methylated arginines

PHD Finger

Possible domain in reader protein that binds methylated lysines, sometimes acetylated lysines

SANT

Possible domain in reader protein that binds unmodified histone tails

HDAC

Histone de-acetylase

HDM

Histone de-methylase

HDAC, HMC, phosphatase

Eraser proteins include:

serine, threonine, histidine, tyrosine

____ and ___ (less commonly ___ and ____) of histone tails can be phosphorylated

H3K9me, H3K27me (Hox silencing and X inactivation)

Gene silencing histone modifications include:

H3K4me, H3K9ac, H3S10ph, H3K14ac

Activating histone modifications include:

Mig1 unbound/Gal4 bound, Mig1 bound (recruits Tup1 which recruits HDAC, reduces Gal4 expression)

In the Gal4/UAS system, the permissive state is ______ while the repressed state is ______

Deacetylation

_____ of the FLC locus triggers flowering

TAF1, Swi/Snf

Proteins that have bromodomains include:

HP1, polycomb

Proteins that have chromodomains include:

TAF1

Histone acetyltransferase containing two bromodomains — needs to bind two adjacent acetyl groups ~7-8 amino acids apart

chromatin condensation, S10phos

HP1 binding promotes _____, ____ prompts HP1 dissociation

H3K4me1, H3K4me3, H3K36me3 (repressive, recruits HDAC)

Active enhancers are marked by _____, active promoters are marked by _____, actively elongated (post-pausing) genes are marked by _____

H3K4me2 and 3/H3K27ac, mostly H3K4me2 and 3, mostly H3K4me2 and 3/H3K27ac

Type 1 promoters are marked with ______, type II are marked with _____, and type III are marked with _____

Paused

Can resume transcription

Arrested

Involves RNAP II backtracking and removal of the 3’ overhang by TFIIS before transcription resumes

Poised

Indicates RNAP II is near the TSS

Stalled

RNAP II has halted transcription

NELF

Pausing transcription factor that associates with the paused/arrested transcription complex and dissociates upon termination

DSIF

Pausing transcription factor that associates with the paused/arrested transcription complex and dissociates upon termination

DRB

Elongation blocker that inhibits P-TEFb and is associated with the arrested transcription complex

P-TEFb

Positive transcription elongation factor b; kinase that phosphorylates RNAP II and DSIF causing NELF to dissociate

Tar sequence, stem loop, host promoter, RNAP II

HIV-provided TAT protein binds _____ of truncated HIV transcripts which forms a _____ that TAT binds. The vicinity of TAT to the ____ increases the rate of transcription initiation because it then binds host _____

P-TEFb, RNAP II from pausing

Recruitment of ____ via TAT releases _____ which strongly increases HIV gene transcription