Eukaryotic transcription factors

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What's a transcription factor?
any protien, other than RNA polymerase, required to initiate or regulate transcription in eukaryotic cells.
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Two types of factors
General - formation of TIC. Specific - transcription of particular genes by binding their regulatory sequences
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Two subdivisions of specific TFs
ubiquitous factors, present in all tissues, and tissue specific inducible factors, synthesised at specific times and tissues
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What do insulators do?
Prevent interference between transcriptional units.
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Structure of a typical pol II promotor
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How do Tf's bind to DNA
Specific amino acid side chain:base interactions. typically 10-20 contacts
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Specific TFs can act with the transcription complex...
directly or indirectly
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Where do you find enhancers and how long are they?
up to 50kbp upstream or downstream of promoter. 50bp-1.5kbp
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Position and consensus sequence for the CAAT box, and factors that bind it.
GGCCAATCT, -80, CTF family, CP1 (ubiq), C/EBP (liver)
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Position and consensus sequence for the GC box, and factors that bind it.
GGGCGG. often multiple sites, -40 to -80. Sp1 binds.
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In what tissue does Oct-2 bind to the octamer sequence?
brain and lymphoid tissue
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Example of the binding sites that Globin genes contain
binding sites for ubiq. factors Sp1 and CP1. and also for tissue specific factors such as GATA-1
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Binding sites that Growth hormone gene contains
2 for Pit-1, 2 for glucocorticoid receptor, 2 for CREB
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What happens if Pit-1 is mutated?
No growth hormone - dwarf phenotype
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What kind of techniques can be used to detect TFs binding DNA
Footprinting techniques
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What does ChIP do
Chromatin immunoprecipitation - detects which DNA sequences are bound by specific tfs in vivo.
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Explain how ChiP works
Protein and DNA are bonded and sheared - a specific protien antibody is used to immunoprecipitate the protien of interest. the DNA sequences that are bound to this protien can be sequenced
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Another way you can show which DNA sequences are important for transcriptional regulation
Mutation experiments combined with reporter gene assay
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How can we purify transcription factors?
affinity chromatography- using their property of binding specific DNA sequences
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The four structural motifs of transcription factors
helix-turn-helix. Zinc finger. Leucine zipper. Helix-loop-helix (homeobox domains)
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How does helix-loop-helix interact with DNA
The C-terminal recognition sequence makes contact with the major groove of DNA.
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Examples of HTH tf's
tryptophan repressor, lambda Cro, lambda repressor fragment, CAP fragment
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What is the homeodomain protien?
a transcriptional regulator that enhances the expression of homeotic selector genes, important in fly development
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Describe the structure of a homeodomain protein
3 a helixes that are tightly folded together by hydrophobic interactions. Helix 2 and 3 resembles the hth with helix 3 being the recognition helix
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Other tf's with homeodomains
POU homeodomain proteins - pituitary specific octamer uncordianted 86.
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What does POU need for dna binding?
Homeodomain and POU specific HTH domain.
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An example of the paired Helix-turn-helix DNA binding domains
Pax6, which also contains a homeodomain
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What do mutations in Pax6 and pax6 do
Pax genes are involved in cell fate during development. Pax6 = aniridia. Pax-3 = waardenburg syndrome type i
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What is common to all zinc finger motifs?
Contain one or more atoms of zinc as structural motifs
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Define type 1 zinc finger
finger loop of 12 amino acids, formed by 2 cys, 2 hys, binding a single zinc atom. Zinc holds an alpha helix and beta sheet together
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How do type 1 zinc fingers interact with DNA
Found in clusters with the alpha helix of each finger interacting with the major groove of DNA
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An example of type 1 zinc finger proteins
Sp1, and GATA factors which are important in embryonic and blood development.
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Why would we re-engineer zinc fingers?
target specific genome sequences for gene therapy. eg, preventing the transcription of HIV receptors.
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Define type 2 zinc fingers
They are nuclear receptors that have two cys2/cys2 fingers, one for DNA binding and the other for dimerisation
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Ligands for type 2 zinc fingers?
Steroids, retinoids, vitamin D, thyroid hormones.
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How do group 1 steroid receptors bind?
Bind as monomers or homodimers to related 6-bp sequences. Arranged as two palindromic repeats
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What effect does cortisol binding have on glucocorticoid zinc finger receptors
allows them to enter the nucleus after the hsp protien has been lost
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Examples of group 2 nuclear receptors - zinc fingers
rar- retinoic acid. T3R, thyroid hormone. VDR, vitamin D.
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How to group 2 nuclear receptors bind?
Bind direct repeat sequences as heterodimers with RXR, which binds 9-cis-retinoic acid.
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What determines what parnters with RXR?
The distane between the direct repeat. eg,, +2 = RAR.
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Other cards in this set

Card 2


Two types of factors


General - formation of TIC. Specific - transcription of particular genes by binding their regulatory sequences

Card 3


Two subdivisions of specific TFs


Preview of the front of card 3

Card 4


What do insulators do?


Preview of the front of card 4

Card 5


Structure of a typical pol II promotor


Preview of the front of card 5
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