AP Biology

Eukaryotic Gene

Expression

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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?

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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

turn genes on & off rapidly flexibility & reversibility

adjust levels of enzymes for synthesis & digestion

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Evolution of gene regulation

Eukaryotes

multicellular

evolved to maintain constant internal conditions while facing changing external conditions homeostasis

regulate body as a whole growth & development

long term processes

specialization turn on & off large number of genes

must coordinate the body as a whole rather than serve the needs of individual cells

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Eukaryotic Gene Expression

Two features of eukaryotic genomes

are a major information-processing

challenge:

First, the typical eukaryotic genome is

much larger than that of a prokaryotic

cell

Second, cell specialization limits the

expression of many genes to specific

cells

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Points of control

The control of gene

expression can occur at any

step in the pathway from

gene to functional protein

1. packing/unpacking DNA

2. transcription

3. mRNA processing

4. mRNA transport

5. translation

6. protein processing

7. protein degradation

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How do you fit all

that DNA into

nucleus?

DNA coiling &

folding

double helix

nucleosomes

chromatin fiber

looped

domains

chromosome

from DNA double helix to

condensed chromosome

1. DNA packing

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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

molecules

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Histones and the Cell Cycle

The association of DNA and histones

seems to remain intact throughout the

cell cycle

This maintains cell identity

DNA is coiled more during mitosis

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Differential Gene Expression

A multicellular organism’s cells

undergo cell differentiation:

specialization in form and function

Differences between cell types result

from differential gene expression

the expression of different genes by

cells within the same genome

In each type of differentiated cell, a

unique subset of genes is expressed

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DNA packing as gene control

Degree of packing of DNA regulates transcription

tightly wrapped around histones

no transcription

genes turned off

Chemical modifications to histones and DNA

of chromatin influence both chromatin

structure and gene

expression

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Histone Modification

In histone acetylation, acetyl groups

are attached to histone tails

This process seems to loosen

chromatin structure, thereby promoting

the initiation of transcription

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Histone Acetylation Acetylation of histones unwinds DNA

loosely wrapped around histones

enables transcription

genes turned on

attachment of acetyl groups (–COCH3) to histones

conformational change in histone proteins

transcription factors have easier access to genes

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DNA Methylation

DNA methylation, the addition of methyl

groups to certain bases in DNA, is

associated with reduced transcription

in some species

DNA methylation causes long-term

inactivation of genes in cellular

differentiation

In genomic imprinting, methylation

turns off either the maternal or paternal

alleles of certain genes at the start of

development

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DNA methylation Methylation of DNA blocks transcription factors

no transcription

genes turned off

attachment of methyl groups (–CH3) to cytosine C = cytosine

nearly permanent inactivation of genes ex. inactivated mammalian X chromosome = Barr body

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Epigenetic Inheritance

Although the chromatin modifications

just discussed do not alter DNA

sequence, they may be passed to future

generations of cells

The inheritance of traits transmitted by

mechanisms not directly involving the

nucleotide sequence is called

epigenetic inheritance

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Eukaryotic Expression

Modeling

Objective: Model how eukaryotic gene expression is controlled.

Your first model should include:

DNA, mRNA, histones (nucleosomes), acetyl groups (for histone acetylation), methyl groups (for DNA methylation), RNA polymerase

Please call me over to evaluate your models when ready. Allmembers of the group must be able to describe what is occurring in each model.

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Regulation of Transcription Initiation

Chromatin-modifying enzymes provide

initial control of gene expression by

making a region of DNA either more or

less able to bind the transcription

machinery

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Organization of a Typical Eukaryotic Gene

Associated with most eukaryotic genes

are regulatory regions called control

elements

segments of noncoding DNA that help

regulate transcription by binding certain

proteins

Control elements and the proteins they

bind are critical to the precise

regulation of gene expression in

different cell types

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The Roles of Transcription Factors

To initiate transcription, eukaryotic

RNA polymerase requires the

assistance of proteins called

transcription factors

An activator is a protein that binds to an

enhancer and stimulates transcription

of a gene

Some transcription factors function as

repressors, inhibiting expression of a

particular gene

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2. Transcription initiation

Control regions on DNA

promoter nearby control sequence on DNA

binding of RNA polymerase & transcription factors

“base” rate of transcription

enhancer distant control

sequences on DNA

binding of activator proteins

“enhanced” rate (high level) of transcription

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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

Turning on Gene movie

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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

Enhancer Sitesregulatory sites on DNA distant from gene

Initiation Complex at Promoter Site binding site of RNA polymerase

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Mechanisms of Post-

Transcriptional Regulation Transcription alone does not account

for gene expression

More and more examples are being

found of regulatory mechanisms that

operate at various stages after

transcription

Such mechanisms allow a cell to fine-

tune gene expression rapidly in

response to environmental changes

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RNA Processing

In alternative RNA splicing, different

mRNA molecules are produced from

the same primary transcript.

Exons may be skipped, shuffled around

or occasionally introns can be included

in the final transcript

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3. Post-transcriptional control

Alternative RNA splicing

variable processing of exons creates a

family of proteins

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4. Regulation of mRNA degradation

Life span of mRNA determines amount

of protein synthesis

mRNA can last from hours to weeks

RNA processing movie

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RNA interference

Small interfering RNAs (siRNA)

short segments of RNA (21-28 bases) bind to mRNA

create sections of double-stranded mRNA

“death” tag for mRNA triggers degradation of mRNA

cause gene “silencing” post-transcriptional control

turns off gene = no protein produced

siRNA

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Action of siRNA

siRNA

double-stranded

miRNA + siRNA

mRNA degradedfunctionally

turns gene off

mRNA for translation

breakdownenzyme(RISC)

dicerenzyme

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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

Control of translation movie

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6-7. Protein processing & degradation

Protein processing

folding, cleaving, adding sugar groups, targeting for transport

Protein degradation

Protein processing movie

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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

7 protein processing & degradation

4

4

Gene Regulation

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Eukaryotic Expression

Modeling

Your first model should include:

DNA, mRNA, histones (nucleosomes), acetyl groups (for histone acetylation), methyl groups (for DNA methylation), RNA polymerase

Your second model should include:

DNA, mRNA, RNA polymerase, promoter, control elements, gene, general transcription factors, activators , enhancers, (transcription initiation complex). You may include other aspects of the transcription initiation complex if you wish.