Post on 11-Jul-2020
Transcription and chromatin
General Transcription Factors +Promoter-specific factors +
Co-activators
Cofactor or Coactivator1. work with DNA specific transcription factors to
make them more effective - examples A. TFIID B. yeast mediator - contains 20 polypeptides
binds to the CTD of RNA polymerase IIC. CBP/p300 works with CREB proteinD. TRAPS - steroid receptor protein cofactorE. OCA-B with Oct 1 proteinF. SAGA complex - contains Ada proteins
(adaptor protein) one of which is also known as Gcn5; works with acidic activators such as Gal4 protein or Gal4-VP16
Cofactor or Coactivator
2. Several of these proteins modify chromatin; acetylation of histone tails
A. TAFII 250B. Gcn5 is one of the first
characterized histone acetyl transferases(HAT)
C. CBP/p300
Gaining Access to the DNA
Large multi-subunit protein complexes alter the state of
chromatin
Organization of Chromatin
Gary Felsenfeld & Mark Groudine, Nature, 2003; 421:448-53.
Schalch, T et. al., Nature. 2005;436:138-41.
Proposed 30nm fiber structure
What is chromatin remodeling?
The process of making DNA more or less accessible in the eukaryotic genome using
a series of specialized proteins
Chromatin Remodeling • ATP-dependent chromatin remodeling
– SWI/SNF and RSC– ISWI: CHRAC, NURF, ACF– CHD/Mi-2– INO80 and SWR1
• Covalent modification– ACETYLATION– METHYLATION– PHOSPHORYLATION– UBIQUITINATION– Poly ADP RIBOSYLATION– SUMOLYATION
What is the functional role of chromatin remodeling
• Global regulation of chromatin structure• Transcription activation and repression• Transcription elongation and termination• DNA repair • DNA recombination
– including immunoglobulin gene rearrangement
• DNA replication
ATP-dependent chromatin remodeling machines
A. they alter the structure of the nucleosome i. the nucleosomal DNA is made more
accessible for binding to transcription factors ii. changes can also be observed by measuring
access to cutting by restriction endonucleasesiii. changes have also been observed in DNA
supercoiling suggesting a change in the path of DNA around the nucleosome
iv. electron microscopy studies have suggested a reduction on the amount of DNA wrapped in the remodeled nucleosome
ATP-dependent chromatin remodeling machines
b. Slides nucleosomes along DNA, changes their translational position
i. changes in electrophoretic mobility is observedii. micrococcal nuclease mapping also shows
changes in translational positioningc. Spaces and assembles nucleosomes
Sliding Nucleosomes
Spacing nucleosomes
Nucleosomes are randomly distributed along DNA
Located equal distances from each other (uniformly spaced, but no uniquely positioned)
These complexes can be recruited to specific chromatin sites by:
A. transcriptional activators such as SWI/SNF by Gal4, Gcn5 and the glucocorticoid receptor protein
B. repressors such as the ISWI complex from yeast called ISW2 by the Ume6 repressor protein
Accessibility is Key Regulatory Step
ATP-dependent chromatin remodeling machinesDifferent classes of chromatin
remodeling complexes
Different classes of ATP-dependent chromatin remodeling complexes
A. SWI/SNF or SWI-SNF like complexes– required for activation of a small subset of
genes– found in humans, flies, and yeast– at least two different forms of the complex in
yeast or humans– in yeast it has 11 or 17 subunits– total complex size is 1.5 -2 Megadaltons
Wiley-Liss, Inc.
Mammalian SWI/SNF
Different classes of ATP-dependent chromatin remodeling complexes
B. ISWI or imitation SWI complexes-its DNA dependent ATPase subunit is
similar to the ATPase subunit of SWI/SNF-has the distinguishing protein domain called
SANT-smaller complexes: only 1-4 subunits-more abundant and play a more global role-are involved in transcriptional repression- there are three distinct ISWI complexes in
flies and yeast
ISWI Subfamily of Chromatin Remodeling Complexes
Corona, D.F. and J.W. Tamkun, Multiple roles for ISWI in transcription, chromosome organization and DNA replication.Biochim Biophys Acta, 2004. 1677(1-3): p. 113-9.
Remodeling Activities of ISWI Subfamily
Langst, G. and P.B. Becker, Nucleosome mobilization and positioning by ISWI-containing chromatin-remodeling factors.J Cell Sci, 2001. 114: p. 2561-8.
Different classes of ATP-dependent chromatin remodeling complexes
C. Mi-2 or CHD complexes-these chromatin remodeling complexes are
associated with histone deacetylases (HDAC) - ATPase subunit has sequence homology to
the ATPase subunit of SWI/SNF belongs to the Swi2 superfamily, but makes up its own subfamily
Different classes of ATP-dependent chromatin remodeling complexes
D. INO80 complex-involved in DNA repair-has DNA helicase activity associated
with the complexE. SWR1 complex
-promotes the exchange of H2A from the nucleosome
-brings in a H2A variant called H2AZ
SNF2
ISWI
Mi-2
INO80
Helicase Bromo AT Hook SANT
Chromo PHD
Chromatin Remodeling Complexes Are Classified by the ATPase Subunits
HAND
SLIDE DBINO
Mechanisms of ATP-dependent chromatin remodeling
Several models involving DNA bulges or DNA twist
Flaus and Owen-Hughes Biopolymers 2003
Twist diffusion model
Would move nucleosomes in 1 bp increments
Least disruptive of histone-DNA contacts
Bulge Propagation Model
Flaus and Owen-Hughes Biopolymers 2003
Propagation of bulge in a wave-like manner around the nucleosome
Planar bulge
• B:\Blaine\Presentations\movies\bulging.mov
• ..\..\Presentations\movies\worming.mov