Transcript Structure of the DNA-binding motifs of activators

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 zinccontaining 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 activatorbinding 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
•
•
•
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.