Prokaryotic RNAP independently bind to their target DNA promoters
T/F
True
Eukaryotic RNAPs independently bind to their target DNA promoters
T/F
False
Unlike prokaryotic RNAP, eukaryotic RNAPs do not independently bind to their target DNA, but are recruited to the promoters through their physical association with Transcription Factors (TFs)
What are the two main groups of promoters for a typical eukaryotic class II gene?
Core promoters
proximal promoter elements/upstream regulatory elements
what are the three main regulatory regions in multicellular eukaryotes?
core promoter
proximal promoter elements
long-range regulatory elements
What does the core promoter of a gene serve as a recognition site for?
It serves as a recognition site for RNAP II and general TF
What happens to the function of a gene if the core promoter is inverted or otherwise moved through a mutation process?
It stops working
Core promoter sequences are only active if positioned in the direct orientation and are inactivated if moved or inverted
Name the classical core promoter elements
TATA box
Initiator element (Inr)
TFIIB recognition element (BRE)
Downstream promoter element (DPE)
Motif ten element (MTE)
not all promoters have all elements
Fill in the blanks
Which sequences of the core promoters are recognized by TFIID?
TATA box
Initiator element (Inr)
Downstream promoter element (DPE)
Motif ten element (MTE)
Which sequences of the core promoters are recognized by TFIID?
BRE (TFIIB recognition element)
What is another name for the TATA box?
Hogness box
How might you characterize the TATA/Hogness box?
an element that can function by itself/ can function in absence of all other core elements
Which subunit of TFIID targets the TATA box?
TBP
(TATA-binding protein)
INR can function independently of TATA
T/F
True
Inr can function independently of TATA, but if present alongside with TATA, the two elements act synergistically to initiate transcription
Which element functions in TATA-less promoters?
DPE
requires presence of Inr element
Which subunits of TFIID recognize the Inr element that spans the transcription start site?
TAF1 and TAF2
(TBP-assocaited factor 1 and 2)
Which subunits of TFIID recognize the DPE element, about 30 bp downstream of the start site?
TAF6 and TAF9
Where is the MTE element located?
(motif ten element)
+18 to +27 downstream of transcription initiation
Which two other elements show strong synergistic effect on transcription when in the presence of MTE?
TATA and DPE
What does MTE require to promote transcription?
the function of Inr
(but TATA and DPE are not required)
What are examples of long-range regulatory elements?
enhancers, silencers, insulators, Locus control regions (LCRs) and matrix attachment regions (MARs)
LCRs
Locus control regions
MARs
Matrix attachment region
Three characteristics of enhancers/silencers and their ability to activate/repress transcription:
At a distance
in either orientation
in either position (upstream, downstream or within a transcription unit)
euchromatin
It represents the major genes and is involved in transcription.
heterochromatin
It mostly consists of repetitive DNA sequences and non-coding RNA transcripts and is relatively gene poor.
What are Insulators
Insulators are 0.3-2kb DNA elements that serve as boundary markers between regions of gene-rich euchromatin and gene-poor condensed heterochromatin
How can Insulators allow for differential regulation of neighboring genes?
It can block enhancer/silencer activity from afar
Locus control regions (LCRs)
DNA sequences that organize and maintain a functional domain of active chromatin and enhance transcription of downstream genes
Classical enhancers are orientation dependent
T/F
False
LCRs are orientation dependent
T/F
True
also, LCRs can enhance expression of several linked downstream genes
Where are MARs typically located
near enhancers
they are thought to confer tissue and stage specific expression to genes in the loops
Matrix attachment regions (MARs)
DNA elements that organize the genome into multiple independent chromatin loops
What GTFs are necessary and sufficient for accurate transcription initiation in vitro?
RNAP II
Six “classical” GTFs: TFIIA, TFIIB, TFIID, TFIIE, TFIIF, and TFIIH
Mediator, a highly conserved, large multi-subunit complex that was originally identified in yeast and is required for transcription initiation in vivo
Describe this image:
Formation of the preinitiation complex by GTFs
TFIID with the help of TFIIA binds the TATA box and forms the DA complex
TFIIB binds next forming the DAB complex
TFIIF binds RNAPII and the two join the DAB complex to form DABPolF complex
The DNase protected footprint changes from -42/-17 in DAB to -42/+17 in DABPolF
What are the last two GTFs to join the pre-initiation complex?
TFIIE and TFIIH
Describe the TBP (TATA-box binding protein)
TBP harbors a very conserved sequence of amino acids that pair with the tata box
Which protein involved in transcription initiation interacts with DNA in the minor groove?
TBP
TBP is a DNA-binding protein that interacts with the DNA minor groove forcing it to open and bending the DNA 80 degrees
TBP only functions with RNAPII promoters that contain TATA
T/F
False
TBP functions not only with RNAPII promoters that contain TATA, but also with TATA-less promoters recognized by RNAPI, II or III!
Describe TBP-associated factors
They interact with the promoter elements and with gene-specific transcription factors
