Genetics Chp 17 Regulation of Gene Expression in Eukaryotes
2 major types of protein are involved in modifying chromatin structure:
1. ATP-dependent chromatin-remodeling complexes 2. Enzymes causing acetylation (addition of CH3COOH groups) and methylation (addition of CH3 groups) of histones.
Harold Weintraub and Mark Groudine hypothesis
Actively transcribing genes are "open genes"
transcription factors
Collection of proteins that mediate the binding of RNA polymerase and the initiation of transcription.
what is chromatin?
DNA + associated Histone proteins
Harold Weintraub and Mark Groudine prediction
IF Actively transcribing genes are "open genes" THEN we should see that actively transcribing genes will be sensitive to DNase, but genes not actively being transcribed will not
RISC
RNA induced silencing complex -degrade mRNA -inhibit translation
Methylation of DNA is correlated with...
gene inactivation
Scientists Harold Weintraub and Mark Groudine looked to see if...
genes they knew were actively transcribing would be more sensitive to DNase
coactivators are also called...
mediators
Proks or Euks: Amount of genetic information
more in eukaryotes
positive control must be at work to...
open up DNA at promoter regions for gene expression to occur.
histone charge
positive
What are snRNPs?
small nuclear ribonucleoproteins that consist of snRNA and proteins
what do coactivators do?
tie basal TF to activators
why is DNA coiled around histones?
to save space
can Chromatin Modifications Can Be Passed to Daughter Cell?
yes
can promoters have varying combinations of regulatory sequences?
yes
Splicing using snRNPs is a stepwise process using snRNPS 1-6. Describe the binding of each snRNP in sequential order.
- U2 binds to branching point of intron - U1 binds to 5' branching point - U3, U4, U5, U6 join U1 - U1 and U4 leave; U5 and U6 mark end on RNA - U2 attacks 5' end of intron (cut) - U2 goes to U5/U6 - U2/U6 leave, stuck on intron - U5 welds gap
name the regulatory tfs
-PPE -silencers -enhancers
Basal TFs
-Proteins that bind to the core promoter near the start point of transcription -Required to recruit RNA polymerase
activators
-Proteins that turn operons on by binding to DNA -Bind PPE or Enhancers
step 1 of transcription initiation
-Regulatory transcription factors bind to DNA -recruit HATs/ATP-dependent chromatin-remodeling complexes -unwind DNA from histone
types regegulatry tfs
-activators -repressors
step 4 of transcription initiation
-basal transcription factors bind core promoter elements (form the basal transcription complex) -recruits RNA polymerase II
How are enhancers and promoter proximal elements different than core promoter regions such as TATA box and BRE?
-core promoter regions are required for all genes -PPE are unique to specific genes
Name the Motifs of TF DNA-binding Domains
-helix turn helix -zinc-finger -leucine zipper
step 2 of transcription initiation
-loosening of the chromatin structure -exposing the promoter region
Describe how these miRNAs carry out their impact on gene expression at both the transcriptional as well as translational level using the terms RITS and RISC.
-miRNA processed in dicer gene --> give double stranded RNA -RITS- interferes with transcription complementary to RNA sequence -RISC- silences expression by binding to RNA; degrades or prevents translation/transcription
repressors
-proteins that block transcription -Bind Silencers
step 3 of transcription initiation
-regulatory TFs bind enhancers and pep -interact with basal transcription factors that bind the promoter
reasons why is Eukaryotic Gene Expression more complex than Prokaryotic
1. More genetic information 2. DNA associated w/ histones. 3. Multiple chromosomes 4. Transcription and translation spatially distinct. 5. 3 RNA polymerases. 6. mRNA processing. 7. needs to be stabalized. 8. Multicellular - many different cell types. 9. Eukaryotes undergo development.
Basal TFs are assembled at promoter in a specific order, which is:
1. TFIID (TAF+TBP) --> TATA box 2. TFIIA & TFIIB --> TFIID 3. RNA pol II joins. 4. Promoter clearance
ways that ATP-dependent chromatin-remodeling complexes work
1. alteration of DNA-histone contacts 2. allegation of DNA path 3. remodeling of nucleosome core particles
Levels of Regulation in Eukaryotes
1. changes in chromatin/chromatin remodeling 2. initiation of transcription 3. RNA processing 4. mRNA stability 5. Post transcriptional protein modification
all of our somatic cells have all the same DNA, yet we KNOW that each tissue type expresses its own set of genes. name some reasons for this
1. chromatin modification can be passed down 2. regulatory TFs- read/silence genes differently for certain cells 3. cell receptors- change external signals, growth, activate/stop TFs 4. splicing
regulation of mRNA at the level of translation includes...
RITS and RISC
RITS
RNA-induced transcriptional silencer -chromatin remodeling silences/strops transcription
where are enhancer located?
Upstream, downstream, or within gene up to 1 million bp away
Enhanceosome
a large protein complex that acts synergistically to activate transcription
Trans-acting regulatory proteins
able to identify and bind target regulatory sequences on any chromosome and change gene activity
example of DNA Modification and its impact:
acetylation- addition of CH3COOH that increases transcription by activating genes
what happens in unregulated transcription?
activator TFs bind enhancer, alter configuration of chromatin
what did Weintraub and Groudine conclude?
actively transcribing genes ARE more unwound from histones than genes that are not actively being transcribed
what does acetalyation do?
adds negative charge to histones, leading to decreased affinity of histones and DNA
Why did they choose to study the B-globin and ovalbumin genes?
b-globulin expressed in blood cells ovalbumin expressed in egg cells only
ribozymes
catalytic RNA molecules that function as enzymes and can splice RNA
where do β-globin and ovalbumin genes come from?
chicken blood cells
are enhancer cis-acting or trans-acting?
cis-acting
basal transcriptions factors are bound to regions of the promoter called...
conserved regions
in these yeast cells, scientists saw...
constitutive expression of genes that were not normally expressed
Proks or Euks: Location of transcription and translation
cytoplasm and simultaneous in prokaryotes
core promoter
determines the accurate initiation of transcription by RNA polymerase II
why is it important that Chromatin Modifications Can Be Passed to Daughter Cell?
differences in modifications can cause different genes to be expressed in different cell
Methylation of histones can be correlated with...
either gene activation or gene inactivation
what are enhancers that up regulate genes called?
enhancers
Proks or Euks: DNA associated histone proteins
eukaryotes
Proks or Euks: Post-transcriptional processing of mRNA
euks have splicing, tail, cap
Proks or Euks: Different cell types requiring different gene expression
euks only
Proks or Euks: Different developmental requirements for gene expression
euks only
Proks or Euks: Number of RNA polymerases
euks-3, proks-1
constitutive
expressed all the time
Acetylation of histones is usually associated with...
gene activation, increase in transcription
Transcriptionally active DNA means it is:
going through transcription
what do regulatory tfs do?
help with recognition, transcription
what is DNA coiled around?
histones
example of Histone modification and its impact:
methylation- addition of CH3 that either increases/decreases transcription of genes
Evidence that Coiling of DNA Inhibits Transcription in brewers yeast
mutant brewer's yeast cell lacked their normal set of histone proteins
microRNAs
naturally occurring small pieces of RNA that can alter gene expression
DNA charge
negative
Results suggest a mechanism of...
negative control
is ovalbumin expressed in blood cells?
no, only in eggs
Results suggest a mechanism of negative control where...
normal default state of eukaryotic genes is to be turned off
Epigenetic inheritance
patterns of inheritance that are not due to differences in gene sequences.
this Also suggests that a form of...
postive control
What type of RNA uses snRNPs and what type has self-splicing rybozymes?
pre-RNA, snRNA
Histone code hypothesis
precise patterns of chemical modifications of histones contain information that influences whether or not a particular gene is expressed
group 1 ribozyme introns
primary transcript of rRNAs.
(PABP) polyadenylate binding protein
protects polyA tail from normal degradation
what does the basal trancription complex do?
recruits RNA polymerase II so that transcription can begin
Promoter-proximal elements
regulatory sequences within 200 bp of promoter that are unique to specific genes, providing a mechanism for eukaryotic cells to exert precise control over transcription
cis-acting regulatory sequences
sections of DNA that are involved in controlling the activity of genes on the same chromosome
group 2 ribozyme introns
self excision by introns in the primary transcripts of mt & cp mRNAs & tRNAs
what are motifs?
shapes
enhancers
short (5-10 bp) DNA sequences that are also involved in gene regulation
AREs (Adenine-Uracil Rich Elements)
signal for rapid turn over; removal of polyA tail
what are enhancers that down regulate genes called?
silencers
snRNA
small nuclear RNA
snRNA is complexed with proteins to form...
small nuclear ribonucleoproteins (snRNP)
snRNP
small nuclear ribonucleoproteins; recognize the splice sites in the pre-mRNA
What is the difference between self splicing ribozymes and introns that use snRNPs?
snRNP- cut out introns and exons self-splicing- cutting introns and exons w/o snRNPs
Spliceosomes are composed of _____.
snRNPs and other proteins
Most gene regulation occurs at...
the initiation of transcription.
Leucine zippers, zinc fingers and helix turn helix motifs all refer to what?
transcription factors/DNA binding sites
t/f Biologists hypothesized that the lack of histone proteins prevents assembly of normal chromatin
true
t/f enhancer cna Work in either orientation
true
Open DNA means it is:
uncoiled
does Pattern of chemical modifications on histones varies from one cell type to another?
yes
is β-globin expressed in blood cells?
yes
which gene was much more cut up by DNase?
β-globin
what type of genes did Weintraub and Groudine examine?
β-globin and ovalbumin genes
