Differential Gene Expression• Transcriptional control

• RNA processing control

• Translational control

• Post-translational control

2

Transcriptional control DNA-mRNA

mRNA processing and stability

Translational control mRNA -Protein

Post-Translational control Protein modifications

3

Promoter Transcription initiation site Translation initiation site

• 5’ UTR (untranslatedsequence)

Exons (expressed) Introns (intervening) Translation termination codon 3’UTR

• Polyadenylation addition site

Transcription termination site

4

5

Transcription initiation site

Translation initiation site

Translation termination codon

3’ Untranslated region

Promoters

Exons

Introns

Three main regulatory agents:• Nuclear proteins that regulate access to DNA

• Multiple, alternative promoters

• Cells differ in number or type of transcription

factors

Let’s take a look at each…..

6

Nuclear proteins regulate access to DNA

DNA normally bound up in nucleosomes (histone proteins)

Chromatin remodeling proteins unwind specific DNA sequences allowing access to transcription initiation complex

• Exposes enhancer and promoter regions allowing transcription

Detour -- now we need to learn what enhancers & promoters are……

7

Chromatin remodeling affects• DNA replication & repair

• Apoptosis

• Chromosome segregation

• Pluropotency

Targeting chromatin remodeling pathways

is a growing field in targeting several

cancers (regulate mitosis)

Promoter -- initiates transcription of a particular gene (TATA box, GC islands)

Enhancer -- short region of DNA which can be found with proteins (TF’s) to “enhance” transcription levels in a gene cluster

Transcription factor -- works in conjunction with enhancers; family of proteins; alter DNA confirmation

Silencer -- “negative” enhancers; DNA sequence which binds repressors preventing RNA polymerase from initiating transcription

Know this

Direct methylation of DNA

Methyltransferases –enzymes that add a methyl group

Common in CpG sequences• “Cytosine – phosphate-Guanine”

In mammals, 70 – 80% of CpG sites are methylated

Repression mechanism:• Blocks transcription factors

Enhancers:• Specific sequence in DNA• Short series of bases• Vary in location• Can be far away from gene regulated • Bound by transcriptional activators (molecules from

inside or outside the cell that aid in transcription)

Silencers• Specific sequence of DNA• Short series of bases• Bind proteins that promote assembly of large protein

complexes that block transcription

11

Know qualities

Many enhancers cause DNA looping• Transcriptional activator

binds to enhancer

• Activator also interacts with part of transcription complex to recruit complex to gene by bending DNA

• Complex then interacts with promoter and recruits polymerase holenzyme

12

Know this

Enhancers are hard to find because:• Variable in sequence from one gene to the next

• Short series of bases

• Vary in location

• Can be far away from gene regulated

So -- how do you find them?

13

“Reporter Constructs”• Put region of interest (with

regulatory elements) upstream

of an inducible gene (such as

lacZ)

• Modify (by deletion or

substitution) presumed

regulatory region

• Modifications that alter

expression of inducible gene

are regulatory elements

14

mRNA processing results in same mature mRNA

Alternative promoters allow for different regulation of same gene• Larval vs. adult expression

Fetal gamma-globin to adult beta-globin in humans

15

Some genes have more than one promoter sequence

Different promoters produce different primary

transcripts

3 major domains:• DNA binding domain

• Activating domain – interacts with initiation complex proteins

• Protein-protein interaction domain

Major transcription factor families• Homeodomain – Hox genes (axis formation)

• Basic helix-loop-helix – muscle & nerve specification

• Basic leucine zipper – liver & fat cell differentiation

– Zinc Finger - segmentation, secondary sex determination, metamorphosis, limb development

16

Know this