BIOMG 1350 - Lecture 18

Gene Expression Regulation I

Gene Expression

The process by which the information encoded in a DNA sequence is translated into a product that has some effect on a cell or organism.

How is gene expression regulated?

Differential Gene Expression Theory

Differential Gene Expression Theory

• The Genome is constant in all somatic cell

• Only a subset of the genome is expressed in any cell type.

• Unused genes that are not transcribed still retain the potential to be expressed

• Different cell types express different genes

Transcriptional Control

• relatively slow acting

• Most energy efficient

• Most common control point for long term regulation such as in development

Inactive portien

• Fast acting

• readily reversible

Prokaryotic Transcription Step 1

RNA polymerase binds to the promoter (stretch of DNA containing certain sequences) with the help of ‘transcription factor(s)’

Prokaryotic Transcription Step 2

Promoter defines where RNA polymerase starts to synthesize RNA

Prokaryotic Transcription Step 3

RNA polymerase synthesizes RNA in 5’ to 3’ direction using the template strand of DNA

Prokaryotic Transcription Step 4

RNA polymerase stops at the terminator (specific DNA sequences), falls off from the DNA and releases the completed RNA

Prokaryotes Model

Are useful model systems for understanding regulation of gene expression

Promoters

Tell RNA polymerase where to start transcription

• a region of DNA that contains sequences recognized and bound by RNA polymerase to start transcription

• Located upstream of the transcription start site

Terminators

Tell RNA polymerase where to stop transcription

• A region of DNA that contains sequences recognized by RNA polymerase as a signal to terminate transcription

• Sequence of terminator is included in the RNA transcription

How are different genes expressed at different levels?

• promoters vary in their strength of binding to RNA polymerase

• promoter activity is enhanced by transcriptional activators

• promoter activity is inhibited by transcriptional repressors

• activators and repressors can act in concert to provide highly sensitive transcriptional regulation

Transcriptional regulators (activators and repressors)

• Are proteins that have DNA binding domains

• Depending on the exact DNA sequence, the transcriptional regulator will bind tightly, loosely, or not at all

• Both prokaryotes and eukaryotes use transcriptional regulators

Operon

Cluster of bacterial genes transcribed from single promoter

• Within the operon’s promoter is an operator, a DNA sequences that is recognized by one or more transcriptional regulators

Operator

A “master switch” for transcription of the genes in the operon

• express all of them or none of them

• specific to prokaryotes

Transcription of the Lac operon genes

Allow E.coli to utilize lactose when glucose is absent

• Lac is only expressed when glucose is present and lactose is present

Lac operon

provides a simple example of gene regulation by transcriptional activators and repressors

Lac repressor

Inhibits transcriptions when lactose isn’t present

Lactose is present

allolactose is a metabolite of lactose, its levels reflect lactose levels

Allolactose

Unbinds Lac repressor from DNA, however inhibiting repressor binding is not sufficient to allow transcription because Lac promoter is weak

CAP

Activates transcription when glucose is not present

CAP Mechanism

CAP binds upstream of the promoter and will only bind when glucose is scarce.

cAMP and Glucose

When glucose levels are low, cAMP levels go up - part of the stress response

cAMP binds to CAP activator

helps to recruit RNA polymerase to the promoter

Glucose And Lactose are Present

• Lactase: Repressor is bound to allolactose and is not bound to DNA

• Glucose: CAP is not bound to cAMP

Operon Off

Glucose present but lactose isn’t

• CAP is unbound

• Repressor bound

• Operon off

Glucose and Lactose aren’t present

• Both CAP and Repressor will be bonded

• Operon off

Glucose not present, lactose present

• CAP will be bounded and allolactose will bind to repressor so receptor will not be bound

• Operon on

Regulation of Transcription in Eukaryotes

• More complex compared to prokaryotes

• Eukaryotes contain 3 RNA polymerases: I,II,III

RNA Pol I

Transcribes rRNA to make ribosomes

RNA Pol II

Transcribes all mRNAs - this will be our focus

RNA Pol III

Transcribes tRNAs for translation

Promoter in Eukaryotics

Contains a TATA Box bound by the TATA Box Binding Protein or TBP, which helps recruit general transcription factors

Results in RNA Pol II

• Collection of proteins that gather at promotor are much more numerous.

• Open the DNA double helix in preparation for transcription

Eukaryotes Enhancers

Act at a distance to regulate gene expression

• Regulatory DNA sequences are bound by transcriptional regulators, which usually (not always) bind activators

• In development, enhancers allow the transcription of a given gene in specific cell types or at specific developmental stages

• Can be located either close to or far away (up to tens of thousands of nucleotides) from the transcription start site

DNA Looping

Allows enhancer to be near the transcription start site even when far away

Enhancer Providing Context Specific Gene Regulation

Gene A only wanted in brain in limb

• To control the gene, enhancers will be used

• Proteins specific to the brain can bind to one of the enhancers and activate it’s expression.

• Limb-expressed transcription factors can bind to enhancer which will allow expression of Gene A

Expression of Shh Changed

• This will cause a change in the enhancer sequence so that it binds to other transcription regulators and will change the expression pattern of the Shh

• One nucleotide change: Extra fingers

Shh Limb Enhancer Experiment

• Substitute the mouse enhancer with the snake enhancer

  • Mouse with no legs

• Make snake enhancer with human enhancer

  • Limb development resorted

How does regulation of gene expression drive development?

Effector is an activator for gene expression, expression of gene will codes a protein that can do other things