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Structure of the DNA-binding motifs of activators

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					Structure of the DNA-binding
     motifs of activators
          Chapter 12
        Categories of Activators
• Activators can stimulate or inhibit
  transcription by RNA polymerase II

• Structure is composed of at least 2 functional
  domains
  – DNA-binding domain
  – Transcription-activation domain
  – Many also have a dimerization domain
        DNA-binding domains

• DNA-binding domains have DNA-binding
  motif
  – Part of the domain having characteristic shape
    specialized for specific DNA binding

  – Most DNA-binding motifs fall into 3 classes
       Zinc-containing modules
• There are at least 3 kinds of zinc-containing
  modules that act as DNA-binding motifs
• All use one or more zinc ions to create a
  shape to fit an α-helix of the motif into the
  DNA major groove
  – Zinc fingers – TFIIIA and Sp1
  – Zinc modules – Glucocorticoid receptor
  – Modules containing 2 zinc and 6 cysteines –
    GAL4
            Homeodomains
• These domains contain about 60 amino acids
• Resemble the helix-turn-helix proteins in
  structure and function
• Found in a variety of activators
• Originally identified in homeobox proteins
  regulating fruit fly development
        bZIP and bHLH Motifs
• A number of transcription factors have a
  highly basic DNA-binding motif linked to
  protein dimerization motifs
  – Leucine zippers
  – Helix-loop-helix
• Examples include:
  – CCAAT/enhancer-binding protein
  – MyoD protein
 Transcription-Activation Domains

• Acidic domains: GAL4

• Glutamine-rich domains: Sp1

• Proline-rich domains: CTF

• Structure and function – not clearly related
           The GAL4 Protein

• Yeast activator controls a set of genes
  responsible for metabolism of galactose

• The GAL4 protein is a member of the zinc-
  containing family of DNA-binding proteins
            Nuclear receptor
• Zinc module - nuclear receptor

• This type of protein interacts with a variety of
  endocrine-signaling molecules

• Protein plus endocrine molecule forms a
  complex that functions as an activator by
  binding to hormone response elements and
  stimulating transcription of associated genes
    Type I Nuclear Receptors
• These receptors
  reside in the
  cytoplasm bound to
  another protein
• When receptors bind
  to their hormone
  ligands:
  – Release their
    cytoplasmic protein
    partners
  – Move to nucleus
  – Bind to enhancers
  – Act as activators
Types II and III Nuclear Receptors
• Type II nuclear receptors stay within the
  nucleus
- Bound to target DNA sites
- Without ligands the receptors repress gene
  activity
- Bind ligands - they activate transcription
• Type III receptors - ligands are not yet
  identified
        Functions of Activators
• Bacterial core RNA polymerase is incapable of
  initiating meaningful transcription

• RNA polymerase holoenzyme can catalyze
  basal level transcription
  – Often insufficient at weak promoters
  – Cells have activators to boost basal transcription to
    higher level in a process called recruitment
         Eukaryotic Activators
• Eukaryotic activators also recruit RNA
  polymerase to promoters
• Stimulate binding of general transcription
  factors and RNA polymerase to a promoter
• 2 hypotheses for recruitment:
  – General TF cause a stepwise build-up of
    preinitiation complex
  – General TF and other proteins are already bound to
    polymerase in a complex called RNA polymerase
    holoenzyme
Models for Recruitment
    Interaction Among Activators
• General transcription factors must interact to
  form the preinitiation complex
• Activators and general transcription factors
  also interact
• Activators usually interact with one another in
  activating a gene
  – Individual factors interact to form a protein dimer
    facilitating binding to a single DNA target site
  – Specific factors bound to different DNA target
    sites can collaborate in activating a gene
              Action at a Distance
• Bacterial and eukaryotic enhancers stimulate
  transcription even though located some distance
  from their promoters
• Four hypotheses attempt to explain the ability
  of enhancers to act at a distance (homework)
  –   Change in topology
  –   Sliding
  –   Looping
  –   Facilitated tracking
Hypotheses of Enhancer Action
           Complex Enhancers
• Many genes can have more than one activator-
  binding site permitting them to respond to
  multiple stimuli

• Each of the activators that bind at these sites
  must be able to interact with the preinitiation
  complex assembling at the promoter - by
  looping out any intervening DNA
          Control Region of the
          Metallothionine Gene




• Gene product helps eukaryotes cope with heavy metal
  poisoning
• Turned on by several different agents
Architectural Transcription Factors


Architectural transcription factors are those
transcription factors - change the shape control
region so that other proteins can interact
successfully to stimulate transcription
            Enhanceosome
• An enhanceosome is a
  complex of enhancer
  DNA with activators
  contacting this DNA

• An example is the HMG
  that helps to bend DNA
  so that it may interact
  with other proteins
    Examples of Architectural
      Transcription Factors

• Besides LEF-1, HMG I(Y) plays a similar
  role in the human interferon-b control gene

• For the IFN-b enhancer, activation seems
  to require cooperative binding of several
  activators, including HMG I(Y) to form an
  enhanceosome with a specific shape
                Homework

• Explain the four hypotheses of the ability of
  enhancers to act at a distance.

• What are insulators? Explain the functions of
  insulators. Explain mechanism of insulator
  activity.
         Study material for exam
•   Structure of activator
•   Three types of DNA binding motif
•   Three types of transcription activation domain
•   What is enhancer? Two types – architectural factors
    and enhanceosome
•   Recruitment
•   Insulator
•   Ubiquitylation
•   Sumoylation
      Regulation of Transcription
               Factors
• Phosphorylation of activators can allow them to
  interact with coactivators that in turn stimulate
  transcription
• Ubiquitylation of transcription factors can mark them
  for
   – Destruction by proteolysis
   – Stimulation of activity
• Sumoylation is the attachment of the polypeptide
  SUMO which can target for incorporation into
  compartments of the nucleus
• Methylation and acetylation can modulate activity
             Ubiquitylation

• Ubiquitylation - monoubiquitylation of some
  activators can have an activating effect

• Polyubiquitylation marks these same proteins
  for destruction
        Activator Sumoylation
• Sumoylation is the addition of one or more
  copies of the 101-amino acid polypeptide
  SUMO (Small Ubiquitin-Related Modifier) to
  lysine residues on a protein
• Process is similar to ubiquitylation
• Results quite different – sumoylated
  activators are targeted to a specific nuclear
  compartment that keeps them stable
         Activator Acetylation
• Nonhistone activators and repressors can be
  acetylated by HATs

• HAT is the enzyme histone acetyltransferase
  which can act on nonhistone activators and
  repressors

• Such acetylation can have either positive or
  negative effects
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