Download Histone modifications

Griffiths • Wessler • Carroll • Doebley
Introduction to
Genetic Analysis
TENTH EDITION
CHAPTER 12
Regulation of Gene Expression
in Eukaryotes
© 2012 W. H. Freeman and Company
CHAPTER OUTLINE
12.1
12.2
12.3
12.4
12.5
Transcriptional regulation in eukaryotes: an overview
Lessons from yeast: the GAL system
Dynamic chromatin
Short-term activation of genes in a chromatin environment
Long-term inactivation of genes in a chromatin
environment
12.6 Gender-specific silencing of genes and whole
chromosomes
12.7 Post-transcriptional gene repression by miRNAs
The first cloned mammal
Dolly and Bonnie
Dolly, the Finn Dorset lamb in 1996
and her surrogate Scottish Blackface mother
Ian Wilmut
Dolly in Royal Museum of Scotland
Dolly
Dolly Parton
Cells differ in gene expression
All cells have the same genome,
but each cell expresses only a subset of all genes
globin
myosin
housekeeping
erythrocyte
muscle cell
All genes
Overview of transcriptional regulation
nucleus
(membrane)
chromatin
Gene regulation at multiple levels
Many regulatory
proteins have to
import into nucleus
Transport
Localization
Modification
Complex formation
Degradation
Promoter-proximal elements precede the promoter of a
eukaryotic gene
Promoter-proximal elements are necessary for efficient
transcription
Point mutations throughout the promoter region were analyzed for their
effects on transcription rates. The height of each line represents the
transcription level relative to a wild-type promoter or promoter-proximal
element (1.0).
Transcription factors need multiple functional domains
1. A domain that recognizes a DNA regulatory sequence
(the protein’s DNA-binding site)
2. A domain that interacts with one or more proteins of the
transcriptional apparatus (RNA polymerase or a protein
associated with RNA polymerase)
3. A domain that interacts with proteins bound to nearby
regulatory sequences on DNA such that they can act
cooperatively to regulate transcription
4. A domain that influences chromatin condensation either
directly or indirectly
5. A domain that acts as a sensor of physiological
conditions within the cell
Direct or indirect (by interacting with other proteins)
Model Organism Yeast
Saccharomyces cerevisiae
Budding yeast
Brewer’s yeast
Baker’s yeast
The Gal pathway
Expressed at low level
Induced by galactose
Regulated by Gal4
Transcriptional activator proteins bind to UAS elements in
yeast
UAS: Upstream Activation Sequences
Binding site for Gal4
Can be far from promoter
Transcriptional activator proteins are modular
Reporter gene
domain swap
Transcriptional activator proteins may be activated by an
inducer
Galactose: inducing signal
Transcriptional activator proteins recruit the transcriptional
machinery
Enhancer can function upstream or
downstream, even far away
Enhancer can function
far away from promoter
Specific recognition
of target sequence
Co-activator:
Does not directly bind DNA
Transcriptional complexes
Combinations of regulatory proteins control cell types
Mating type
Combinations of binding partners => different binding specificities
The structure of chromatin
~150 bp
linker DNA
A nucleosome is composed of DNA wrapped around eight
histones
Histone octamer (H2A2H2B2H32H42)
DNA exposed on the outside
The structure of chromatin
The structure of chromatin
Euchromatin
(loose)
Heterochromatin
•Condensed
•Repetitive sequences
•Late replicating
•Genes silenced
Chromatin remodeling exposes regulatory sequences
Linker DNA: sensitive to nuclease
Nucleosomal DNA: protected from nuclease digestion
Use nuclease sensitivity to determine chromatin state
(open/closed) or nucleosome position
+ SWI-SNF + ATP
Shifting of nucleosome position
Exposes regulatory sequences
The SWI-SNF complex for chromatin remodeling
Yeast mutant screen
sugar nonfermenting (snf)
Mating type switch (swi)
swi2=snf2
swi2/snf2 (“switch-sniff”) locus
SWI-SNF complex
Modifications of histone tails results in chromatin remodeling
Histone tails are exposed, can be modified
Modifies Lysine (K) and Arginine (R) (basic aa)
Acetylation: negative charges => repulsion
Histone modifications
Histone modifications
Acetylation of histones
Histone acetyltransferase (HAT)
Histone de-acetylase (HDAC)
Histone modifications
Histone code
Alternative modifications on
the same residue
Regulation of gene expression by histone acetylation
Histone deacetylation can turn off gene transcription
HDAC
(corepressor)
Inheritance of chromatin states
Epigenetic memory: heritable traits (over rounds of cell division and
sometimes transgenerationally) that do not involve changes to the
underlying DNA sequence. (e.g. chromatin state)
Methylation of DNA
A model for the inheritance of DNA methylation
In mammals, 70-80% of CG are methylated genome-wide.
CpG island: clusters around gene promoter
maintenance
hemi-methylated
Enhanceosomes help recruit the transcriptional machinery
Enhanceosomes recruit chromatin remodelers
Enhancers contain binding sites for
many transcription factors, which bind
and interact cooperatively.
Enhancer-blocking insulators prevent enhancer activation
Model for how enhancer-blocking insulators might work
Mating-type switching is controlled by recombination of DNA
cassettes
silent information regulators (SIR)
Sir2 (HDAC)
ds break in MAT made by HO
endonuclease
=> gene conversion
Gene silencing is caused by the spread of heterochromatin
Position-effect variegation (PEV)
w+ is expressed in some cells => not a mutation in w gene
Clonal => epigenetic memory
Heterochromatin in Drosophila chromosomes
~30% of genome
H3K4me2, enriched in euchromatin
H3K9me2, enriched in heterochromatin
Some genes enhance or suppress the spread of
heterochromatin
Enhancer
Suppresor
Su(var)2-5 = HP1 (heterochromatic protein 1)
Su(var)3-9 = histone methylatransferase
Multiple states of Lysine methylation
Heterochromatin may spread farther in some cells than in
others
Barrier insulators stop the spread of heterochromatin
Genomic imprinting
Genomic imprinting
Phenotype depends on the parental origin of the genes
Inactivation of genes and chrosomomes
Genomic imprinting
mouse/human
~100 imprinted genes
No changes in DNA sequence
Genomic imprinting requires insulators
imprinting control region
DNA methylation
Unusual inheritance of imprinted genes
Steps required for imprinting
Igf2: maternal imprinting (inactive)
H19: paternal imprinting (inactive)
Igf2 H19
X inactivation
Dosage compensation for X chromosome
female: XX
male: XY
Barr body and Lyon Hypothesis of X inactivation
Murray Barr: discoverer
Mary Lyon
Epigenetic memory
Xi: H3K9me, histone hypoacetylation, DNA hypermethylation
~ heterochromatin
Xist
non-coding
Xist RNA covers one of the two copies of the X chromosome
RNA fluorescent in situ hybridization (FISH)
metaphase chromosomes
female fibroblast cell line
Xist expression => cis-inactivation
A model for X-chromosome inactivation
Possible models for the repression of translation by miRNA