Chapter 18 Regulation of Gene Expression

Classify these things as occurring in prokaryotes, eukaryotes, or both. Single loop of DNA Chromosomes wound around histones Telomeres Multiple sites of origin in DNA replication Single origin site for DNA Replication Uses DNA polymerase RNA is processed after transcription Transcription/translation can be coupled Translation occurs at the ribosome Uses codons and anticodons

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.

Gene Structures

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.

trp Operon Makes Tryptophan. Repressible 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.

Wednesday, February 18 Predict what would happen in the lac operon for each of these scenarios. Lactose is present, glucose is scarce A mutation in the operator so the repressor cannot bind Lactose is absent Glucose is present The repressor has a mutation so that it always binds to the operator CAP and cAMP levels are high

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 (-COCH 3 ) to AAs in histones. Result - DNA held less tightly to the nucleosomes, more accessible for transcription.

DNA Methylation Addition of methyl groups (-CH 3 ) 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 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-X L – inhibits apoptosis Bcl-X S – induces apoptosis Two different and opposite effects!!

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 translation. 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 homeotic genes? 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.

Monday, February 23 During mitotic cell division, each daughter cell receives an exact copy of the DNA from the parent cell. Explain two mechanisms how eukaryotes have exactly the same copy of DNA in each cell, yet different proteins can be expressed.

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 (common example) 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, proto-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.