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