Transcript Chromatin Structure and Its Effects on Transcription

Chromatin Structure and Its Effects
on Transcription
Chapter 13
Histones
• Eukaryotic cells contain 5
kinds of histones
–
–
–
–
–
H1
H2A
H2B
H3
H4
• Each histone type - not
homogenous
– Gene reiteration
– Posttranslational
modification
Features of Histones
• Abundant proteins
• Most are well-conserved from one species to
another
• Not single copy genes - repeated many times
– Some copies are identical
– Others are quite different
– H4 has - 2 variants
Nucleosomes
• Chromosomes are long, thin molecules –tangle - if
not carefully folded
• Folding occurs in several ways
- First order of folding is the nucleosome
– Maurice Wilkins - X-ray diffraction has shown strong
repeats of structure at 100Å intervals
– 110Å spacing
Histones in the Nucleosome
• Chemical cross-linking in solution: (Korenberg)
– H3 to H4
– H2A to H2B
• H3 and H4 exist as a tetramer (H3-H4)2
• Chromatin is composed of roughly equal
masses of DNA and histones
– 1 histone octamer/200 bp of DNA
– Octamer composed of:
• 2 each of H2A, H2B, H3, H4
• 1 each of H1
H1 and Nucleosomes
• Treatment of chromatin with trypsin or high salt
buffer removes histone H1
• Left chromatin looking like “beads-on-a-string”
• The beads named nucleosomes
– Core histones form a ball with DNA wrapped around the
outside
– H1 also lies on the outside of the nucleosome
Histone Acetylation
• Histone acetylation occurs in both cytoplasm and nucleus
• Cytoplasmic acetylation carried out by HAT B (histone
acetyltransferase - HAT)
– Prepares histones for incorporation into nucleosomes
– Acetyl groups later removed in nucleus by deacetylases
Histone Acetylation
• Nuclear acetylation of core histone N-terminal tails
– Catalyzed by HAT A
– Attracts bromodomain proteins - essential for
transcription
– Correlates with transcription activation
– Coactivators of HAT A found which may allow loosening
of association between nucleosomes and gene’s control
region
Histone Deacetylation
• Transcription repressors bind to DNA sites
and interact with corepressors which in turn
bind to histone deacetylases
– Repressors
• Unliganded nuclear receptors
• Mad-Max
– Corepressors
• NCoR/SMRT
• SIN3
– Histone deacetylases - HDAC1 and 2
Ternary Protein Complexes
• Assembly of complex
brings histone
deacetylases close to
nucleosomes
• Deacetylation of core
histones allows
– Histone basic tails to bind
strongly to DNA+ histones
in neighboring
nucleosomes
– This inhibits transcription
Activation and Repression
Source: Adapted from Wolfe, A.P., 1997. Sinful repression. Nature 387:16-17.
Deacetylation of core histones removes binding sites
for bromodomain proteins that are essential for
transcription activation
Chromatin Remodeling
• Activation of many eukaryotic genes requires
chromatin remodeling
• Several protein complexes carry this out
– All have ATPase harvesting energy from ATP
hydrolysis for use in remodeling
– Remodeling complexes are distinguished by
ATPase component
Remodeling Complexes
• SWI/SNF
– In mammals, has BRG1 as ATPase
– 9-12 BRG1-associated factors (BAFs)
• A highly conserved BAF is called BAF 155 or 170
• Has a SANT domain responsible for histone binding
• This helps SWI/SNF bind nucleosomes
• ISWI
– Have a SANT domain
– Also have SLIDE domain involved in DNA
binding
SWI/SNF Chromatin Remodeling
Mechanism of Chromatin
Remodeling
• Mechanism of chromatin remodeling involves:
– Mobilization of nucleosomes
– Loosening of association between DNA and core histones
• Catalyzed remodeling of nucleosomes involves
formation of distinct conformations of nucleosomal
DNA/core histones when contrasted with:
– Uncatalyzed DNA exposure in nucleosomes
– Simple nucleosome sliding along a DNA stretch
Heterochromatin
• Euchromatin: relatively extended and open
chromatin that is potentially active
• Heterochromatin: very condensed with its
DNA inaccessible
– Microscopically appears as clumps in higher
eukaryotes
– Repressive character able to silence genes as much
as 3 kb away
Heterochromatin and Silencing
• Formation of at tips of yeast chromosomes
(telomeres) with silencing of the genes is the
telomere position effect (TPE)
• Depends on binding of proteins
– RAP1 to telomeric DNA
– Recruitment of proteins in this order:
• SIR3
• SIR4
• SIR2
SIR Proteins
• Heterochromatin at other locations in
chromosome also depends on the SIR proteins
• SIR3 and SIR4 interact directly with histones
H3 and H4 in nucleosomes
– Acetylation of Lys 16 on H4 in nucleosomes
prevents interaction with SIR3
– Blocks heterochromatin formation
• Histone acetylation also works in this way to
promote gene activity
Histone Methylation
• Methylation of Lys 9 in N-terminal tail of H3
attracts HP1
• This recruits a histone methyltransferase
– Methylates Lys 9 on a neighboring nucleosome
– Propagates the repressed, heterochromatic state
• Methylation of Lys and Arg side chains in
core histones can have either repressive or
activating effects
Modification Interactions
• The modifications
shown above the tail are
activating
– Ser phosphorylation
– Lys acetylation
• Modification below the
tail (Lys methylations) is
repressive
Modification Combinations
• Methylations occur in a given nucleosome in
combination with other histone modifications:
– Acetylations
– Phosphorylations
– Ubiquitylations
• Each particular combination can send a different
message to the cell about activation or repression of
transcription
• One histone modification can also influence other
nearby modifications
Nucleosomes and Transcription
Elongation
• An important transcription elongation facilitator is
FACT (facilitates chromatin transcription)
– Composed of 2 subunits:
• Spt16
– Binds to H2A-H2B dimers
– Has acid-rich C-terminus essential for these
nucleosome remodeling activities
• SSRP1 binds to H3-H4 tetramers
– Facilitates transcription through a nucleosome by
promoting loss of at least one H2A-H2B dimer from
the nucleosome
• Also acts as a histone chaperone promoting readdition of H2A-H2B dimer to a nucleosome that
has lost such a dimer
•
•
•
This project is funded by a grant awarded under the President’s Community Based Job Training Grant as
implemented by the U.S. Department of Labor’s Employment and Training Administration (CB-15-162-06-60).
NCC is an equal opportunity employer and does not discriminate on the following basis:
against any individual in the United States, on the basis of race, color, religion, sex, national origin, age disability,
political affiliation or belief; and
against any beneficiary of programs financially assisted under Title I of the Workforce Investment Act of 1998
(WIA), on the basis of the beneficiary’s citizenship/status as a lawfully admitted immigrant authorized to work in
the United States, or his or her participation in any WIA Title I-financially assisted program or activity.
Disclaimer
• This workforce solution was funded by a grant awarded under the
President’s Community-Based Job Training Grants as implemented
by the U.S. Department of Labor’s Employment and Training
Administration. The solution was created by the grantee and does
not necessarily reflect the official position of the U.S. Department of
Labor. The Department of Labor makes no guarantees, warranties,
or assurances of any kind, express or implied, with respect to such
information, including any information on linked sites and including,
but not limited to, accuracy of the information or its completeness,
timeliness, usefulness, adequacy, continued availability, or
ownership. This solution is copyrighted by the institution that
created it. Internal use by an organization and/or personal use by
an individual for non-commercial purposes is permissible. All other
uses require the prior authorization of the copyright owner.