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 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. b. Operator c. Structural Genes Gene Structures Regulatory Gene

• Makes Protein which may bind to the operator. • Repressor protein blocks . 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.

• 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%20Ope ron.htm trp Operon

• Makes/synthesizes . • 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

• 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

• Modifications • DNA Methylation • Epigenetic Inheritance Histone Acetylation

• Attachment of acetyl groups (-COCH3) to AAs in . • Result - DNA held less tightly to the , 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

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

• Enhancers and • 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. 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 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. 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.