8. control of gene expression Flashcards
gene expression varies in different cells
- human cells contain ~22,00 genes, but only a fraction of them may be expressed in any one cell
- some genes are expressed in all cells all the time AKA housekeeping genes responsible for routine metabolic functions common to all cells (eg respiration)
- some genes are expressed as cell begins to differentiate
- some genes are expressed only in cells with specialized functions
- some genes are expressed only when surrounding conditions change (arrival of a hormone)
regulatory sequence
stretches of DNA, which through the action of specific proteins, control the activity/expression of genes
what are some major contributing factors that makes us different from chimps?
differences in how the two species regulate the expression of genes
ie. when where and how much the genes are expressed
control of gene expression in eukaryotes terms
transcription level
processing level
translational level
post-translation
transcription level
if and how often a gene is transcribed
transcribed is making RNA using DNA template ( copying the genetic info)
Processing level
different messenger RNAs made from a given gene (alternative splicing)
translational level
how much of the mRNA is made into protein (and mRNA lifetime)
Post-translation
protein lifetime
transcriptional control
Rna polymerase II transcribes some genes much more frequently than others
- this depends on regulatory sites on the DNA and the presence of transcription factors
RNA polymerase is a complex of ______ different proteins
12
transcription factors
DNA binding proteins that regulate transcription
- they have a DNA binding domain
- they have an activation domain (a binding site for other proteins)
- often have a site for binding a second subunit to form a dimer
What are three common motifs in transcription factors
zinc-finger motif
helix-loop helix
leucine-zipper motif
zinc-finger motif
multi-finger shaped projections of protein git into major grooves of DNA
Helix-loop-helix (HLH) motif
forms homo and heterodimers; humans have ~120 HLH genes- can produces 1000s of different transcription factors
Leucine-zipper motif
two helices zipped together to form a coiled-coil structure as dimers
components of the promoter
core promoter
proximal promoter
distal promoter
enhancers
Core promoters
DNA sequence -1 to -40 bases from the start of the coding DNA
- on/off regulation of the gene
- it’s recognized by a series of DNA-binding proteins: general transcription factors, comprising the pre-initiation complex
- core promoter determines whether or not transcription can take place
general transcription factors (GTF)
- TBP (TATA binding protein) -recognizes the TATA sequence about 30 bases upstream from the start of the gene
- TAFs (TBP-associated factors) a group of GTF accessory proteins necessary for the RNA Polymerase II
- RNA polymerase II-produces the RNA
Proximal promoter
-CAAT and GC boxes are bound to transcription factors such as NF1
-NF1 recruits a co-activator needed for RNA polymerase to work
-the proximal elements regulate frequency of transcription
When Cs with GC box are methylated by the cell, the gene is transcriptionally inactive
Distal promoter
contains response elements (-500 to-1000 bp)
- response elements.
- DNA sequence that bind to proteins called specific transcriptions factors
- the cell controls gene activity by regulation the presence or absence of the specific transcription factor
- transcription factors are often activated by dimerization
- every gene has its own set of response elements
Specific transcription factors
may activate or repress transcription acitivty. specific to one gene (or a few genes)
enhancers
- specific DNA sequences, often found far from the target gene, which bind specific transcriptional activators (or enhancer binding proteins), and activate gene expression.
- tens of thousands of bases away;
- can be in any orientation
- on enhancer, when activated, can activate a number of genes
- enhancers are separated from each other by insulator sequences that bind to proteins that separate loops
Glucocorticoid Receptor
transcription factor that controls expression of the PEPCK gene
-structure: DNA binding domain
Activation domain
glucocorticoid Receptor 6 steps
- glucocorticoid (cortisol) is released from adrenal gland following tissue damage
- Cortisol passes through liver cell membrane
- cortisol bind to glucocorticoid receptor-receptor changes shape
- a newly exposed NLS allows entry into nucleus
- glucocorticoid receptor w/ bound hormone can bind to a glucocorticoid response element (GRE)- binds as a dimer-PEPCK gene transcription begins- newly made RNA leaves the nucleus
- PEPCK protein made- gluconeogenesis begins (make glucose to help in stress)
DNA binding domain
Recognizes specific DNA sequence
- since it is a dimer it recognizes a palindromic sequence
Activation domain
Alters transcription, usually through a copressor or coactivator
how does glucocorticoid receptor turn on the gene?
Brings in coactivators that
a) supply general transcription factors for RNA polymerase II, TAFS
b) alter chromatin structure
PEPCK
regulates glucose metabolism
regulated by many different hormones
how do transcription factors affect gene transcription?
by altering histone-binding and making the gene accessible to RNA polymerase
How a trancription factor works?
binds- to the response elements in the distal promoter
regions( recognizes the nucleotide base sequence )
recruits-proteins which help the pre-initiation complex work. called
coactivators
enhances- the RNA polymerase activity
coactivators link transcription factors to:
- general transcription factors needed for transcription
2. chromatin re-modelling enzymes
for the glucocorticoid receptor, the coactivator protein is called:
CBP coactivator, a type of histone acetyltransferase (HAT)
CBP coactivator, a type of histone acetyltransferase (HAT)
acetylates the lysine (K) residues of the histones
has 2 effects:
a) reduces the strength (destabilizes) of the histone-DNA interaction
b)reduces interactions between the histone proteins
histone acetyl transferase step 1
CBP reduces histone binding and TATA box is exposed
histone acetyltransferase step 2
pre initiation of complex has its own histone acetyltransferase activity
- subunit TAFII250 keeps acetylating the histones as it transcribes
- all this is going at about 30 nucleotides per second
transcriptional repression:
histone deacetylases (HDACs)
Histone methyl transferases
DNA methyl transferases
histone deacetylases (DHACs)
remove acetyl groups, and DNA winds up more tightly-gene activity is silences
histone methyltransferases
methyl groups added to histones causes tighter DNA binding- gene activity is silenced
DNA methyltransferases
add methyl groups to DNA- always at carbon 5 of cytosine
this essentially tags regions of DNA so that they are utilized (transcribed differently-usually a sign of inactivation this is a reversible process, but DNA methylation is passed on
