Control of Eukaryotic Genes

(Ch. 19)

The BIG Questions…

• How are genes turned on & off in eukaryotes?

• How do cells with the same genes differentiate to perform completely different, specialized functions?

Evolution of gene regulation

• Prokaryotes– single-celled– evolved to grow & divide rapidly– must respond quickly to changes in

external environment• exploit transient resources

• Gene regulation- Operons– turn genes on & off rapidly

• flexibility & reversibility– adjust levels of enzymes

for synthesis & digestion

Evolution of gene regulation

• Eukaryotes– multicellular– evolved to maintain homeostasis– regulate body as a whole

• specialization–turn on & off large number of

genes• must coordinate the body as a whole

rather than serve the needs of individual cells

Points of control

• The control of gene expression can occur at any step in the pathway from gene to functional protein

from DNA double helix to condensed chromosome

1. DNA packing

Nucleosomes • “Beads on a string”

– 1st level of DNA packing– histone proteins

• 8 protein molecules• positively charged amino acids • bind tightly to negatively charged DNA

8 histone 8 histone moleculesmolecules

DNA packing as gene control• Degree of packing of DNA regulates

transcription– tightly wrapped around histones

• no transcription• genes turned off

heterochromatindarker DNA (H) = tightly packed

euchromatinlighter DNA (E) = loosely packed

DNA methylation

• Methylation of DNA blocks transcription factors – attachment of methyl groups (–CH3) to cytosine

– nearly permanent inactivation of genes• ex. inactivated mammalian X chromosome =

Barr body

Histone acetylation Acetylation of histones

unwinds DNA Loose histone wrapping

enables transcription genes turned on

attachment of acetyl groups (–COCH3) to histones

conformational change in histone proteins

transcription factors have easier access to genes

Chromatin changes

Transcription

RNA processing

mRNA degradation

Translation

Protein processingand degradation

Histonetails

DNAdouble helix Amino acids

availablefor chemicalmodification

(a) Histone tails protrude outward from a nucleosome

(b) Acetylation of histone tails promotes loose chromatinstructure that permits transcription

Unacetylated histones Acetylated histones

2. Transcription initiation• Control regions on DNA

– promoter• nearby control sequence on DNA• binding of RNA polymerase & transcription

factors– enhancer

• distant control sequences on DNA

• binding of activator proteins

Model for Enhancer action

• Enhancer DNA sequences – distant control sequences

• Activator proteins – bind to enhancer sequence &

stimulates transcription

• Silencer proteins – bind to enhancer sequence &

block gene transcription

Transcription complex

Enhancer

ActivatorActivator

Activator

Coactivator

RNA polymerase II

A

B F E

HTFIID

Core promoterand initiation complex

Activator Proteins• regulatory proteins bind to DNA at

distant enhancer sites• increase the rate of transcription

Coding region

T A T A

Enhancer Sitesregulatory sites on DNA distant from gene

Initiation Complex at Promoter Site binding site of RNA polymerase

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Many control elements for different genes are the same

It is the combination of control elements

that provides specificity

3. Post-transcriptional control

• Alternative RNA splicing– variable processing of exons creates a

family of proteins

4. Regulation of mRNA degradation

• Life span of mRNA determines amount of protein synthesis– mRNA can last from hours to weeks

RNA interference NEW!

Chromatin changes

Transcription

RNA processing

mRNAdegradation

Translation

Protein processingand degradation

Degradation of mRNAOR

Blockage of translation

Target mRNA

miRNA

Proteincomplex

Dicer

Hydrogenbond

The micro-RNA (miRNA)precursor foldsback on itself,held togetherby hydrogenbonds.

1

2 An enzymecalled Dicer movesalong the double-stranded RNA, cutting it intoshorter segments.

2

One strand ofeach short double-stranded RNA isdegraded; the otherstrand (miRNA) thenassociates with acomplex of proteins.

3 The boundmiRNA can base-pairwith any targetmRNA that containsthe complementarysequence.

4 The miRNA-proteincomplex prevents geneexpression either bydegrading the targetmRNA or by blockingits translation.

5

A Brief, RNAi Cartoon

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miRNA’s• Scientists discovery video• Animation of miRNA function

RNA interference 1990s | 2006

Andrew FireAndrew FireStanfordStanford

Craig MelloCraig MelloU MassU Mass

“for their discovery of RNA interference —gene silencing by

double-stranded RNA”

5. Control of translation

• Block initiation of translation stage – regulatory proteins attach to 5' end of mRNA

• prevent attachment of ribosomal subunits & initiator tRNA

• block translation of mRNA to protein

6-7. Protein processing & degradation

• Protein processing– folding, cleaving, adding sugar groups,

targeting for transport• Protein degradation

– ubiquitin tagging– proteasome degradation

Ubiquitin

• “Death tag”– mark unwanted proteins with a label – 76 amino acid polypeptide, ubiquitin– labeled proteins are broken down rapidly

in "waste disposers"• proteasomes

1980s | 2004

Aaron CiechanoverAaron CiechanoverIsraelIsrael

Avram HershkoAvram HershkoIsraelIsrael

Irwin RoseIrwin RoseUC RiversideUC Riverside

Proteasome • Protein-degrading “machine”

– cell’s waste disposer– breaks down any proteins

into 7-9 amino acid fragments• cellular recycling

initiation of transcription

1

mRNA splicing

2

mRNA protection3

initiation of translation

6

mRNAprocessing

5

1 & 2. transcription - DNA packing - transcription factors

3 & 4. post-transcription - mRNA processing

- splicing- 5’ cap & poly-A tail- breakdown by siRNA

5. translation - block start of translation

6 & 7. post-translation - protein processing - protein degradation

7protein processing & degradation

4

4

Gene Regulation

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Concept Check

1. Do miRNA’s increase or decrease gene expression?

2. Does histone de-acetylation increase or decrease gene expression?

Genome Sizes and Estimated Numbers of Genes*

 Types of DNA sequences in the human genome

Exons (regions of genes codingfor protein, rRNA, tRNA) (1.5%)

RepetitiveDNA thatincludes

transposableelements

and relatedsequences

(44%)

Introns andregulatorysequences

(24%)

UniquenoncodingDNA (15%)

RepetitiveDNA

unrelated totransposable

elements(about 15%)

Alu elements(10%)

Simple sequenceDNA (3%)

Large-segmentduplications (5–6%)

Just how little of out Genome Codes for Genes?

QuickTime™ and aH.264 decompressor

are needed to see this picture.

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Cancer results from genetic changes in cell

cycle control

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Proto-oncogene --> oncogene

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Multistep Model of cancer

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Genome Evolution

• How did our genome get so big?

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• The similarity in the amino acid sequences of the various globin proteins– Supports this model of gene duplication and

mutation

Table 19.1

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Evolution of Genes with Novel Functions

• The copies of some duplicated genes– Have diverged so much during

evolutionary time that the functions of their encoded proteins are now substantially different

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Concept check

1. How much of the human genome consists of exons?

2. How can exon shuffling lead to the evolution of a new gene

Turn yourQuestion Genes on!

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A larger portion of the DNA in a eukaryotic cell is transcribed than would be predicted by the proteins made by the cell. What is being transcribed and what is its function?

1. Multiple enhancer regions are being transcribed to amplify the transcription of protein-coding genes.

2. These transcriptions are of non-coding “junk” DNA that is a remnant of mutated protein coding segments. The transcripts are degraded by nuclear enzymes.

3. The additional DNA that is transcribed is introns that are excised fro the primary transcript in the produciton of mRNA.

4. Many RNA coding genes are transcribed. Precursor RNAs fold into hairpin structures, which are cut and processed into miRNAs that regulate translation of mRNAs.

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How is the coordinated transcription of genes involved in the same pathway regulated?

1. The genes are transcribed in one transcription unit, although each gene has its own promoter.

2. The genes are located in the same region of the chromosome, and enzymes de-acetylate the entire region so that transcription may begin.

3. A steroid hormone selectively binds to the promoters for all the genes.

4. The genes have the same combination of control elements in the enhancer that bind with the particular activators present in the cell.

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Histones are

1. Small positively charged proteins that bind tightly to DNA

2. Small bodies in the nucleus involved in rRNA synthesis

3. Basic units of DNA packing consisting of DNA wound around a protein core.

4. DNA bending proteins that facilitate formation of transcription initiation complexes.

5. Proteins responsible for producing repeating sequences at telomeres.

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Heterochromatin

1. Has a higher degree of packing than does euchromatin.

2. Is visible with a light microscope during interphase.

3. Is not actively involved in transcription

4. Makes up metaphase chromosomes

5. Is all of the above.

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What is the main reason that prokaryotic genes average 1,000 nucleotride base pairs whereas human genes average about 27.000 base pairs?

1. Prokaryotes have smaller, bue many more individual genes.

2. Prokaryotes are more ancient organisms, longer genes arose later in evolution.

3. Prokaryotes are much simpler organism humans have many types of differentiated cells.

4. Prokaryotic genes do not have introns, human genes have many

5. Human proteins are much larger and more complex than prokaryotic proteins.

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A eukaryotic gene typically has all of the following associated with it except

1. A promoter

2. An operator

3. Enhancers

4. Introns and exons

5. Control elements.