Chapter 18

Regulation of

Gene Expression

Regulation of Gene Expression

• Important for cellular control and differentiation.

• Understanding “expression” is a “hot” area in Biology.

General Mechanisms

1. Regulate Gene Expression

2. Regulate Protein Activity

Operon Model

• Jacob and Monod (1961) - Prokaryotic model of gene control.

• Always on the National AP Biology exam!

Operon Structure

1. Regulatory Gene

2. Operon Area

a. Promoter

b. Operator

c. Structural Genes

Gene Structures

Regulatory Gene

• Makes Repressor Protein which may bind to the operator.

• Repressor protein blocks transcription.

Promoter

• Attachment sequence on the DNA for RNA polymerase to start transcription.

Operator

• The "Switch”, binding site for Repressor Protein.

• If blocked, will not permit RNA polymerase to pass, preventing transcription.

Structural Genes

• Make the enzymes for the metabolic pathway.

Lac Operon

• For digesting Lactose.

• Inducible Operon - only works (on) when the substrate (lactose) is present.

If no Lactose

• Repressor binds to operator.

• Operon is "off”, no transcription, no enzymes made

If Lactose is absent

If Lactose is present

• Repressor binds to Lactose instead of operator.

• Operon is "on”, transcription occurs, enzymes are made.

If Lactose is present

Enzymes

• Digest Lactose.

• When enough Lactose is digested, the Repressor can bind to the operator and switch the Operon "off”.

Net Result

• The cell only makes the Lactose digestive enzymes when the substrate is present, saving time and energy.

Animation

• http://www.biostudio.com/d_%20Lac%20Operon.htm

trp Operon

• Makes/synthesizes Tryptophan.

• Repressible Operon.

– Predict how it is different from the inducible operon…

If no Tryptophan

• Repressor protein is inactive, Operon "on” Tryptophan made.

• “Normal” state for the cell.

Tryptophan absent

If Tryptophan present

• Repressor protein is active, Operon "off”, no transcription, no enzymes

• Result - no Tryptophan made

If Tryptophan present

Repressible Operons

• Are examples of Feedback Inhibition.

• Result - keeps the substrate at a constant level.

Positive Gene Regulation

• Positive increase of the level of transcription.

• Uses CAP - Catabolite Activator Protein

• Uses cAMP as a secondary cell signal.

CAP - Mechanism

• Binds to cAMP.

• Complex binds to the Promoter, helping RNA polymerase with transcription.

Result

• If the amount of glucose is low (as shown by cAMP) and lactose is present, the lac operon can kick into high gear.

Eukaryotic Gene Regulation

• Can occur at any stage between DNA and Protein.

• Be prepared to talk about several mechanisms in some detail.

Chromatin Structure

• Histone Modifications

• DNA Methylation

• Epigenetic Inheritance

Histone Acetylation

• Attachment of acetyl groups (-COCH3) to AAs in histones.

• Result - DNA held less tightly to the nucleosomes, more accessible for transcription.

DNA Methylation

• Addition of methyl groups(-C H3) to DNA bases.

• Result - long-term shut-down of DNA transcription.

• Ex: Barr bodies, genomic imprinting

Epigenetics

• Another example of DNA methylation effecting the control of gene expression.

• Long term control from generation to generation.

• Tends to turn genes “off”.

Do Identical Twins have Identical DNA?

• Yes – at the early stages of their lives.

• Later – methylation patterns change their DNA and they become less alike with age.

Transcriptional Control

• Enhancers and Repressors

• Specific Transcription Factors

• Result – affect the transcription of DNA into mRNA

Enhancers

• Areas of DNA that increase transcription.

• May be widely separated from the gene (usually upstream).

Posttranscriptional Control

• Alternative RNA Processing/Splicing – Ex. - introns and exons

• Can have choices on which exons to keep and which to discard.

• Result – different mRNA and different proteins.

Another Example

Results

• Bcl-XL – inhibits apoptosis

• Bcl-XS – induces apoptosis

• Two different and opposite effects!!

DSCAM Gene

• Found in fruit flies

• Has 100 potential splicing sites.

• Could produce 38,000 different polypeptides

• Many of these polypeptides have been found

Commentary

• Alternative Splicing is going to be a BIG topic in Biology.

• About 60% of genes are estimated to have alternative splicing sites. (way to increase the number of our genes)

• One “gene” does not equal one polypeptide (or RNA).

Other post transcriptional control points

• RNA Transport - moving the mRNA into the cytoplasm.

• RNA Degradation - breaking down old mRNA.

Translation Control

• Regulated by the availability of initiation factors.

• Availability of tRNAs, AAs and other protein synthesis factors. (review Chapter 17).

Protein Processing and Degradation

• Changes to the protein structure after translation.

• Ex: Cleavage– Modifications

– Activation

– Transport

– Degradation

Protein Degradation

• By Proteosomes using Ubiquitin to mark the protein.

Noncoding RNA

• Small RNA molecules that are not translated into protein.

• Whole new area in gene regulation.

• Ex - RNAi

Types of RNA

• MicroRNAs or miRNAs.

• RNA Interference or RNAi using small interfering RNAs or siRNAs.

• Both made from RNA molecule that is “diced” into double stranded (ds) segments.

RNAi

• siRNAs or miRNAs can interact with mRNA and destroy the mRNA or block transcription.

• A high percentage of our DNA produces regulatory RNA.

Morphogenesis

• The generation of body form is a prime example of gene expression control.

• How do cells differentiate from a single celled zygote into a multi-cellular organism?

Clues?

• Some of the clues are already in the egg.

• Cytoplasmic determinants – chemicals in the egg that signal embryo development.

• Made by Maternal genes, not the embryo’s.

Induction

• Cell to cell signaling of neighboring cells gives position and clues to development of the embryo.

Fruit Fly Studies

• Have contributed a great deal of information on how an egg develops into an embryo and the embryo into the adult.

Homeotic (Hox) Genes

• Any of the “master” regulatory genes that control placement of the body parts.

• Usually contain “homeobox” sequences of DNA (180 bases) that are highly conserved between organisms.

Comment

• Evolution is strongly tied to gene regulation. Why?

• What happens if you mutate the homeoticgenes?

• Stay tuned for more “evo-devo” links in the future.

When things go wrong

Example case

• Bicoid (two tailed) – gene that controls the development of a head area in fruit flies.

• Gene produces a protein gradient across the embryo.

Result

• Head area develops where Bicoid protein levels are highest.

• If no bicoid gradient – get two tails.

Other Genes

• Control the development of segments and the other axis of the body.

Gene Expression and Cancer

• Cancer - loss of the genetic control of cell division.

• Balance between growth-stimulating pathway (accelerator) and growth-inhibiting pathway (brakes).

Proto-oncogenes

• Normal genes for cell growth and cell division factors.

• Genetic changes may turn them into oncogenes (cancer genes).

• Ex: Gene Amplification, Translocations, Transpositions, Point Mutations

Proto-oncogenes

Tumor-Suppressor Genes

• Genes that inhibit cell division.

• Ex - p53, p21

Cancer Examples

• RAS - a G protein.

• When mutated, causes an increase in cell division by over-stimulating protein kinases.

• Several mutations known.

Cancer Examples

• p53 - involved with several DNA repair genes and “checking” genes.

• When damaged (e.g. cigarette smoke), can’t inhibit cell division or cause damaged cells to apoptose.

Carcinogens

• Agents that cause cancer.

• Ex: radiation, chemicals

• Most work by altering the DNA, or interfering with control or repair mechanisms.

Multistep Hypothesis

• Cancer is the result of several control mechanisms breaking down (usually).

• Ex: Colorectal Cancer requires 4 to 5 mutations before cancer starts.

Colorectal Cancer

News Flash

• Severe damage to a chromosome that causes it to “shatter” can lead to immediate cancer.

• Doesn’t always take a long time and multiple steps.

Can Cancer be Inherited?

• Cancer is caused by genetic changes but is not inherited.

• However, oncogenes can be inherited.

• Multistep model suggests that this puts a person “closer” to developing cancer.

Example – BRAC1

• BRAC1 is a tumor suppressor gene linked with breast cancer.

• Normal BRAC1 – 2% risk.

• Abnormal BRAC1 – 60% risk.

• Runs in families. Some will have breasts removed to avoid cancer risk.

Summary

• Know Operons

• Be able to discuss several control mechanisms of gene expression.

• Be familiar with gene expression and development of organisms.

• How control of DNA can lead to cancer.