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Regulation of Gene
Activity in
Eucaryotes
1.
2.
3.
4.
Transcription
RNA processing
mRNA transport
mRNA
degradation and
storage
5. Translation
6. Posttranslational
modulation of
protein activity
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Chapter 12. Transcriptional Activators in Eucaryotes
Chapter 11. General Transcription Factors in
Eucaryotes.
Eucaryotic RNA polymerases rely upon TF for
binding to promoters.
For increased level and fine control…
activators..specific TF bind to upstream region of
promoters…enhancers
Eucaryotic cells have gene-specific transcription
factors (activators).
1. To increase transcription,
2. Permits cells to control expression of their genes.
(stimulate or inhibit)
Activators have 2 functional domains
• • Transcriptional-activation domain
• -
DNA-binding Domains
1. Zinc-containing modules. A variety. Use 1> zinc ions
to arrange protein motif so that alpha helix resides
within DNA major groove.
A. B. -)
C. Modules containing 2 zinc and 6 cysteines such
as yeast activator GAL4
2. Homeodomains. 60 aa.
3. Beta barrels. 4. bZIP and bHLH motifs. Highly basic domain and
leucine zipper/HLH
Transcription-activating Domains
1. Acidic domains (acid blob)- GAL4…49 aa domian
with 11 acidic residues
2. Glutamine-rich domains- Sp1- 2 domains of 25%
glutamine.
3. Proline-rich domains- CTF 19/84 aa proline
Structure and importance unclear
Protein/DNA interactions
Specific interactions between bases and aa
Direct
Nondirect (salt
bridges, through
water molecules)
DNA-binding domains
1. Zinc fingers. -
beta/alpha
3D Structure:
Finger contour- dashed
line
Left side of finger is
alpha helix,
-
Zinc finger interaction with DNA
Major Groove.
Curve of protein matches curve of DNA helix. –wrap
around.
Binding between each finger and DNA binding site
relies on direct aa-base interactions
A. aa preceding alpha helix
B. residues 2 and 3 within helix.
Utility of alpha helix contacting major groove.
B- bind DNA backbone and help position alpha
helix
Most of the DNA contacts on one strand. Most
contact bases.
Details:
mouse Zif266immediate early
protein.
-
2. GAL4 protein- yeast activator that controls genes
for metabolism of galactose.
Each gene has a GAL4 target site upstream of start.
Target sites= -.
First 65 aa- DNA binding motif
65-94- similar to zinc finger (zinc and cysteine). But
different…..6 cysteines and no histidines. One zinc
per 3 cysteines.
Recognition: -
3. Nuclear Receptors
Members of this class (-
Ex. Sex hormones, glucocorticoids, Vit D, thyroid
hormone, retinoic acid
A. DNA binding domain with 2 zinc containing
modules. 4 cysteines.
B. Protein protein interaction
GR in inactive form in nucleus complexed with hsp.
Glucocorticoid diffuses across cell membrane and
enters cytoplasm.
Transcriptional
activation.
1. Binding domain
dimerizes, making
contacts with DNA.
2. 3. Binding domain a zinc
module that contains 2
zinc ions.
4. -.
5. Alpha helical interaction
Homeodomains: Members of the HTH family of DNA binding
domains. Each domain contains:
3 alpha helixes: 2 and 3 form HTH motif, 3 is
recognition helix. One helix in major groove, other
across DNA at angle
Engrailed homeodomain. Recognition helix fits into
DNA major groove and makes specific contacts.
Beta- barrels
The factor dimerizes by forming a beta barrel at the
interface between the 2 subunits. Shaped like a barrel,
composed of anti-parallel beta sheets. Positions alpha
helices into major groove.
bZIP and bHLH
DNA binding (b= basic region in each domain that
forms the majority of the DNA binding domain) and
dimerization (leucine zipper and HLH).
Model for Leucine Zipper
2 alpha helices.
Leucines interact like
a zipper. -
bHLH proteins dimerize through a HLH domain which
allows basic residues of each long helix to interact with
the DNA target site.
One face of helix
hydrophobic residues,
other face charged.
Length of connecting loop
12-28 amino acids
Transcriptional Activation Domains:
Stimulate binding of general TF to a promoter.
Build up of complex.
In turn, promotes polymerase binding.