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Gabe Villares

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Enhancesome is a protein complex that binds to an enhancer ... Bending of the enhancer sequence is critical for the formation of an enhancesome ... – PowerPoint PPT presentation

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Title: Gabe Villares


1
TRANSCRIPTION REGULATION
Vidya Gopalakrishnan
MOD.1.017c, MDACC vgopalak_at_mdanderson.org
Cancer Cell Signaling GS040133,  03/18-20/09
2
TOPICS TO BE COVERED
WEB MAKING MODEL
3
COMPLEXITY OF CHROMATIN STRUCTURE
  • Bead on a string structure
  • Compaction
  • Protection
  • But.creates problems for transcriptional and DNA
    replication machinery

Campbell NE et al (Eds) Biology Concepts
Connections 4th Edition, 2003
4
OVERVIEW OF TRANSCRIPTION
Basal Transcription Cis promoters Trans Pol-II
complex Regulation of Transcription Cis
enhancers, silencers Trans activators,
repressors, tissue-specific factors Chromosome
Remodeling Dealing with the chromosome Unpacking
chromatin Histone assembly, Histone modifications
ATP-dependent chromatin remodeling complexes
5
Basal Transcription (Cis-acting elements
promoters)
6
COMPOSITION OF A PROMOTER
Schematic of a typical gene regulatory region.
The promoter, which is composed of a core
promoter and proximal promoter elements,
typically spans less than 1 kb. Distal (upstream)
regulatory elements, which can include enhancers,
silencers, insulators, and locus control regions,
can be located up to 1 Mb pairs from the
promoter. These distal elements may contact the
core promoter or proximal promoter through a
mechanism that involves looping out the
intervening DNA.
7
PROMOTER PROXIMAL ELEMENTS
Core promoter defined as the minimal DNA fragment
that is sufficient to direct correct basal levels
of transcription initiation by Pol II in vitro on
naked DNA templates containing a single
well-defined transcription start site (TSS).
8
UNIVERSALITY OF CORE PROMOTERS?
  • High throughput studies have identified several
    classes of core promoters based on the
    differential usage of the TSS. At the extreme
    ends of the spectrum are two classes of core
    promoter elements Single peak promoters and
    Broad peak promoters
  • Characteristics of Single Peak Promoters
  • Have a relatively tight defined TSS position
    within a few base-pairs
  • More likely associated with TATA boxes
  • Possess a higher frequency of transcription
    factor binding sites
  • TATA-box associated single peak promoters often
    involved in tight regulation of genes, many of
    which are tissue-specific
  • Characteristics of Broad Peak Promoters
  • Show random distribution of many TSSs within a
    100-bp window
  • Often lack TATA boxes and often carry CpG
    islands
  • Possess a lower frequency of transcription
    factor binding sites
  • Often associated with ubiquitously expressed
    genes

9
REDEFINING CORE PROMOTERS
  • TATA box associated with only 10 of promoter
    elements
  • Even when a TATA-box is present, it may not be
    critical for defining promoter activity
  • The function of the TATA box in determining the
    TSS can be attributed to other elements in
    different promoters. None of these elements are
    present in all promoters
  • INR Initiator
  • BRE Transcription factor II B Recognition
    Element
  • DCE Downstream Core Element
  • DPE Downstream Core Promoter Element
  • XCPE1 X-gene Core Promoter Element

Muller and Tora, J Biol Chem. 2007
10
Basal Transcription Trans RNA Pol II and
Associated Factors
11
CLASSIFICATION OF TRANSCRIPTION FACTORS
  • General Transcription factors
  • Examples TFIID, TFIIB, TFIIA, TFIIH, TFIIE,
    TFIIF
  • Expressed in all cell types
  • Ubiquitous transcription factors
  • Examples SP1, CCAAT-box binding protein
  • Expressed ubiquitously, activate transcription of
    many genes
  • Tissue specific transcription factors
  • Examples MyoD, REST, HNF1
  • Bind to specific DNA sequences
  • Express in specific cell types
  • Activate transcription of tissue specific genes

12
MATCHING CORE PROMOTERS WITH RECOGNITION FACTORS
  • Transcription initiation by RNA polymerase II
    involves the assembly of TFIIA, TFIIB, TFIID,
    TFIIE, TFIIF, and TFIIH--in a precise sequence of
    events
  • Binding of the general transcription factor
    TFIID, to the core promoter is the first step in
    the assembly of the whole transcriptional
    machinery
  • TFIID COMPLEX TATA Binding Protein
    (TBP)TBP-Associated Factors (TAFs)
  • TBP may be present at promoters, but may not
    correlate with promoter activation
  • How do TFIID complexes form? A core of TAFs
    (TAF4, 5, 6, 8, 9, 10 and 12) form a minimal
    TFIID complex
  • This minimal TFIID complex can then associate
    with TBP and other TAFs to form a variety of
    TFIID complexes to help stabilize the complex
  • Examples of additional TAFs that recognize
    specific DNA elements
  • TAF1 and TAF2 contact INR
  • TAF1 contact DCE-containing promoters
  • TAF6/TAF9 contact DPE-containing promoters
  • TFIID alone cannot promote efficient DNA binding
    and transcriptionTFIIB, F, E and H required (in
    that order)..critical for PolII binding and
    successful transcription

13
CORE PROMOTERS AND FACTORS THAT BIND THEM
Muller and Tora, J Biol Chem. 2007
14
EXAMPLE OF TATA-ASSOCIATED PROMOTERp21/WAF1
Gartel, A. L. et al. Cancer Res 2005
15
hTert TATA-less promoter
Current Cancer Drug Targets, 2006, Vol. 6, No. 2
hTERT human telomerase reverse transcriptase
  • TERT promoter is GC-rich, lacks TATA and CAAT
    boxes, but contains binding sites for other
    transcription factors
  • Telomerase activity is predominantly regulated at
    the level of transcription

16
HUMAN TELOMERASE IN CANCERS
  • Human telomeres comprise a 2-30 kb array of
    duplex TTAGGG repeats
  • hTERT catalytic subunit of human telomerase
  • Highly active in immortalized cells and more than
    90 of human cancers
  • Is stringently repressed in most normal somatic
    cells

Englert lab webpage
17
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18
ACTIVATOR-DEPENDENT TRANSCRIPTION
  • ACTIVATORS
  • Characterized by distinct DNA binding domains
    such as cysteine-rich zinc finger, homeobox,
    helix-loop helix (HLH), basic helix loop zipper
    (bZIP), forkhead, Pit-Oct-UNC (POU)
  • In addition, they contain a distinct
    transcription activation domain
  • CO-ACTIVATORS
  • Adaptor proteins that typically lack intrinsic
    DNA binding properties, but, provide a link
    between activators and general transcription
    machinery usually in response to a cellular cue

D.B. Nikolov S.K. Burley (1997), PNAS 94, 15-22.
19
OTHER PROMOTER PROXIMAL ELEMENTS AND BINDING
PARTNERS
Transcription factors
Consensus binding site
Specificity protein 1 (Sp1)
GGGCGG
CCAAT/Enhancer binding protein (C/EBP)
CCAAT
Activator protein 1 (AP1)
TGACTCA
Octamer binding proteins
ATGCAAAT
(OCT-1 and OCT-2)
 
E-box binding proteins (E12, E47, E2-2)
CANNTGa
G.M. Cooper, The Cell
20
COMPLEXITIES OF TRANSCRIPTION
Transcriptional activity is greatly stimulated by
activators, which bind to upstream regulatory
elements and work, at least in part, by
stimulating preinitiation complex (PIC) formation
through a mechanism thought to involve direct
interactions with one or more components of the
transcriptional machinery. Activators consist of
a DNA-binding domain (DBD) and a separable
activation domain (AD) that is required for the
activator to stimulate transcription. The direct
targets of activators are largely unknown.
21
Regulation of Basal Transcription Cis Enhancers,
Silencers and Insulators Trans Proteins that
bind to these elements
22
ENHANCER AND DNA LOOPING
  • Enhancer
  • Consists of multiple motifs
  • Binding sites for transcription activators
  • DNA looping
  • Helps bring transcription factors bound at
    distant enhancers to interact with general
    transcription factors at the promoter

G.M. Cooper, The Cell
23
ENHANCER EFFECTS
  • The enhancer is active
  • Upstream or downstream from the transcription
    start site (C and D)
  • In either the forward or backward orientation (E)

G.M. Cooper, The Cell
24
Synergy or additive effect?
ENHANCER ACTIVITY
The increase in transcription rate is higher than
that expected of an additive effect
25
(No Transcript)
26
ENHANCESOME FORMATION
  • Enhancesome is a protein complex that binds to an
    enhancer
  • Best characterized enhancesome is that of the
    human beta-interferon gene that is upregulated in
    response to virus infection
  • cJun/ ATF-2, NF-kappa B and IRF7 bind directly to
    the upstream enhancer proteins upon viral
    infection
  • Interaction mediated by HMG-I, which helps
    stabilize the complex through protein-protein
    interactions
  • Bending of the enhancer sequence is critical for
    the formation of an enhancesome

NFkB
D. Thanos and T. Maniatis, 1995, Cell
27
TRANSCRIPTIONAL SILENCER
  • Block the binding of activators to regulatory
    sequences
  • Block the activation domain of activator
  • Active repression domains inhibit transcription
    by interactions with general transcription
    factors.

G.M. Cooper, The Cell
28
Repressive Activities in the Context of the P21
promoter
Gartel, A. L. et al. Cancer Res 2005
Gartel, A. L. et al. Cancer Res 2005
29
Regulation of Transcription Tissue Specific
Transcription Factors
30
TISSUE-SPECIFIC TRANSCRIPTIONAL REGULATION
Use of alternative TAFs such as the
ovarian-specific TAF105 allows formation of
specialized RNA polymerase initiation complexes
that direct the transcription of tissue-specific
and gene-selective program(s) of expression.
TISSUE SPECIFICITY
Levin and Tjian, Nature, 2003
31
TRANSCRIPTION UNITY IN COMPLEXITY
32
W. R. TAYLOR
Preparation for the upcoming challenge
33
READING RESOURCES
  • Books
  • The Cell A molecular approach, Cooper, Geoffrey
    M.Sunderland (MA) Sinauer Associates, Inc.
    c2000
  • Molecular Biology of the Cell, Alberts, Bruce
    Johnson, Alexander Lewis, Julian Raff, Martin
    Roberts, Keith Walter, Peter Garland Science
    c2002
  • Molecular Cell Biology, Lodish, Harvey Berk,
    Arnold Zipursky, S. Lawrence Matsudaira, Paul
    Baltimore, David Darnell, James E.New York
    W.H.Freeman Co, c1999
  • Molecular biology 2nd edition, Robert.F.Weaver

34
READING RESOURCES (contd)
  • PAPERS
  • Nikolov, DB and Burley, SK. RNA Ploymerase
    transcription initiation - A structural view.
    Proc Natl. Acad. Sci. USA 9415-22 (1997)
  • Orphanides, G and Reinberg, D. RNA Polymerase II
    elongation through chromatin. Nature 407 471-475
    (2000)
  • Tapscott, SJ. The circuitry of a master switch
    MyoD and the regulation of skeletal muscle gene
    transcription. Development. 132 2685-2695 (2005)
  • Szutorisz, H et al. The role of enhancers as
    centres for general transcription factor
    recruitment. Trends in Biochem. Sci. 30593-599
    (2005)
  • Maston, GA et al. Transcriptional regulatory
    elements in the human genome. Ann Rev Human
    Genet. 7 29-59 (2006)
  • Goldberg, AD et al. EpigeneticsA landscape takes
    shape. Cell 128 635-638 (2007)
  • Nagy, Z and Tora, L. Distinct GCN5/PCAF-containing
    complexes function as co-activators and are
    involved in transcription factor and global
    histone acetylation. Oncogene 26 5341-5357
    (2007)
  • Muller, F et al. New problems in RNA Polymerase
    II transcription initiation Matching the
    diversity of core promoters with a variety of
    recognition factors. J. Biol. Chem 282
    14685-14689 (2007)
  • Armstrong, JA. Negotiating the nucleosome-factors
    that allow RNA Polymerase II to elongate through
    chromatin. Biochem. Cell. Biol. 85 426-434
    (2007)
  • Li, B et al. The role of chromatin during
    transcription. Cell 128 707-719 (2007)
  • Kouzarides, T. Chromatin modifications and their
    functions. Cell 128 693-705 (2007)
  • Wang, GG et al. Chromatin remodeling and cancer,
    part1-covalent histone modifications. Trends in
    Mol. Med. 13 363-372 (2007)
  • Ooi, L and Wood, IC. Chromatin crosstalk in
    development and disease lessons from REST.
    Nature Rev. Genet. 8 544-554 (2007)
  • Panning, B and Taatjes, DJ. Transcriptional
    regulation--it takes a village. Mol Cell 31
    622-629 (2008)
  • Suganama, T and Workman, JL. Cross-talk among
    histone modifications. Cell 135 604-607 (2008)
  • Juven-Gershon, T et al. The RNA polymerase II
    core promoter - the gateway to transcription.
    Curr. Opin. Cell. Biol. 20253-259 (2008)
  • Wade, JT and Struhl, K. The transition from
    transcription initiation to elongation. Curr.
    Opin. Genet. Devpt. 18130-136 (2008)
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