Factors for Initiation at Class I Promoters

1
Factors for Initiation at Class I Promoters

• Need nRNAP I and two factors, SL1 and UBF
• SL1 and UBF (a.k.a. IBF1) both bind to both
  elements (UCE Core)
• SL1 provides the species specificity
• SL1 TBP and 3 TAFIs
• TAFIs (110,63, and 48 kDa) different from the
  TAFs in TFIID

2
Initiation at a Class I promoter

• UBF (IBFI UBF) binds to the UCE and core
  promoter.
• SL1 binds to UBF, and the UCE and core elements.
• RNAPI binds at core element.

3
Factors for nRNAP III

• Transcription of 5S rRNA requires 3 factors
  besides nRNAP III
• TFIIIA - binds the internal promoter, a
  Zn2- finger protein
• TFIIIC also binds to the internal promoter
• TFIIIB binds to TFIIIC, enabling TFIIIB to bind
  upstream and recruit the Pol to the transcrip.
  start site

4
Model for the assembly of the pre-initiation
complex on a class-III tRNA promoter.
Note does not need TFIIIA.
Fig. 11.52
5
TFIIIB contains TBP 2 TAFIIIs
In vitro data indicates that TBP used by all 3
polymerases (TFIID, SL1, TFIIIB Fig. 11.54 in
Weaver). In vivo evidence TBP mutant of yeast
did not synthesize 45S pre-rRNA, a mRNA or 5S
rRNA (fIg. 11.11).
6
Pre-initiation complexes on TATA-less promoters
contain TBP.
Assembly factor (green) binds, which promotes
binding of a TBP-complex.
Fig. 11.54
7

• The importance of TBP

TBP important for forming many transcription
initiation complexes it acts by recruiting other
proteins, either RNAPs or other general
transcription factors. The specificity of TBP for
different promoter classes resides with its TAFs.
8
Sp1 Factor for Upstream (Proximal) Class II
Promoter Element

• Binds GC boxes, stimulates transcription
• Interacts with TAFII110 in TFIID
• Also stimulates transcription of TATA-less nRNAP
  II promoter (by promoting TFIID binding)

9
Structure of Eukaryotic Transcription Factors

• Many have modular structure
• DNA-binding domain
• Transcription activating domain
• Proteins can have gt 1 of each, and they can be
  in different positions in protein.
• Many also have a dimerization domain

10
Recent data suggests SP1 actually has 4
activating domains.
11
Activation Domains

• Acidic (e.g., GAL4, 49 aa domain 11 acidic
  aa)
• Glutamine-rich (e.g., 2 in Sp1, 25 gln)
• Proline-rich (e.g., CTF, 84 aa domain 19 are
  proline)

12
DNA-binding domains

• Zinc containing motifs
• Zinc fingers (Sp1 and TFIIIA)
• Zinc modules (GR and other nuclear receptors)
• Modules with 2 Zinc ions and 6 cysteines (GAL4)
• Homeodomains ( 60-aa domains originally found in
  homeotic mutants)
• bZIP and bHLH motifs (a highly basic DNA-binding
  domain and a dimerization domain (leucine zipper
  or helix-loop-helix)

13
Amino acid side chains in proteins can form
H-bonds to DNA bases. Critical for
sequence-specific binding to DNA.
14
3 views of C2H2 Zinc fingers
Often found as repeats in a protein. Bind in the
major groove of DNA.
15
GAL4-DNA Complex

• DNA-binding domain
• 2 Zn2 bound by 6 cysteines
• A Short a helix that docks into major groove

Dimerization domain - Coiled coil (a helices)
Fig. 12.5
16
- Homeotic mutants have wrong organs
(organ-identity mutants) - Occur in animals and
plants - Important regulatory genes
Heres looking at ..uhh..you.
antennapedia
Wild-type
17

• Homeotic genes are transcription factors!
• Have a conserved DNA-binding domain
  (Homeodomain) that resembles a helix-loop-helix
  (HLH) domain.
• Bind DNA as a monomer

18
bZIP proteins

• Have DNA binding and dimerization domains
• DNA binding region is very basic (R and K
  residues)
• Dimerization involves Leucine Zipper
• Can form heterodimers!!

19
Leucine Zipper

• helices form a coiled coil with interdigitating
  leucines.

Fig. 12.12