Gabe Villares

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TRANSCRIPTION REGULATION
Vidya Gopalakrishnan
MOD.1.017c, MDACC vgopalak_at_mdanderson.org
Cancer Cell Signaling GS040133,  03/18-20/09
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TOPICS TO BE COVERED
WEB MAKING MODEL
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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
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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
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Basal Transcription (Cis-acting elements
promoters)
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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.
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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).
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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

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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
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Basal Transcription Trans RNA Pol II and
Associated Factors
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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

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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

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CORE PROMOTERS AND FACTORS THAT BIND THEM
Muller and Tora, J Biol Chem. 2007
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EXAMPLE OF TATA-ASSOCIATED PROMOTERp21/WAF1
Gartel, A. L. et al. Cancer Res 2005
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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

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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
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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.
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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
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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.
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Regulation of Basal Transcription Cis Enhancers,
Silencers and Insulators Trans Proteins that
bind to these elements
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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
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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
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Synergy or additive effect?
ENHANCER ACTIVITY
The increase in transcription rate is higher than
that expected of an additive effect
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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
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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
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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
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Regulation of Transcription Tissue Specific
Transcription Factors
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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
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TRANSCRIPTION UNITY IN COMPLEXITY
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W. R. TAYLOR
Preparation for the upcoming challenge
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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

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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)