Biology 331 Chapter 6 Regulation of Gene Transcription

Biology 331: Chapter 6 Regulation of Gene Transcription

Introduction: n n n Every cell contains all of the genes of that organism How are genes only turned on in the proper setting? Tissue types Changes in the environment Cells must be able to turn transcription on and off Cells must be able to recognize environmental conditions

Prokaryotic transcription control and the lac operon:

Anatomy of the lac operon: The lac operon consists of 6 parts n It contains three genes that deal with lactose metabolism (Z, Y, A) n These genes are transcribed as one m. RNA n

Promoter and operator Up stream of these genes is a promoter region n The promoter is required for RNA polymerase to bind n Between the promoter and the genes is an operator region n

The I gene Up stream of the promoter region is the I gene n The I gene codes for a repressor protein n The repressor protein binds to the operator region n This blocks transcription n

Operon: n. A set of adjacent genes which are transcribed as one unit plus related regulatory sequences

So how does it all work? n If there is no lactose in the environment do I want to produce enzymes to digest lactose? n The I gene constantly produces a repressor which binds to the operator (O) region n Since the action of RNA polymerase is blocked by the repressor transcription does not take place

Lac operon

Allosteric changes

Add lactose to the environment n n n We want to have enzymes to metabolize lactose Lactose binds to an allosteric site on the repressor causing a confirmational change In this case lactose is acting as the "inducer" The repressor no longer binds to the operator Transcription proceeds This is termed negative control since the repressor normally blocks production

Lac operon

Mutations in the system:

The I gene: n Constitutive Mutants: Enzymes are always expressed in an uncontrolled fashion n What happened? n This mutation is recessive. . . explain n

I- Mutations

The Is mutation: n The allosteric site is altered n The protein can no longer bind to the inducer (in this case lactose) n The lactose enzymes are never produced n Explain n This mutation is dominant n Also called trans (across) dominance

Is mutation

The O gene: n Oc mutations: n Changes the sequence of the operator region so the repressor won't bind n Genes on that chromosome are always transcribed n This is a cis (adjacent) dominant mutation n It only affects transcription of genes on the same chromosome n Explain. . .

Oc mutation

The promoter region: n Changes in the promoter region may cause RNA polymerase not to bind n Blocks transcription n Cis Dominant

Catabolite repression of the lac operon: second control system on the lac system n Some energy sources are preferred over others n If glucose AND lactose are present E. coli prefers the glucose n So we must find a way to shut down the lac system in the presence of glucose n This is termed positive control since it only works in the presence of a substance n. A

How does it work? n n n The presence of a glucose metabolite interferes with the production of c. AMP from ATP CAP is produced by the crp gene CAP binds to c. AMP forming a complex This complex binds to the promoter region and increases the affinity of RNA polymerase for the promoter Thus with too much glucose we get too little c. AMP and RNA polymerase affinity is not increased

CAP-c. AMP system

Glucose & No Lactose

Glucose and Lactose

Lactose & no Glucose

Multiple Positive Controls: n The arabinose operon: n n Enzymes are the ara. B, ara. A, and ara. D genes ara. I (initiator) region contains the promoter A product from the ara. C gene binds to arabinose This complex activates transcription

This system also contains the same CAP-c. AMP system already discussed

Positive control

Alternate action of ara. C n ara. C protein binds to both the ara. O and ara. I regions when arabinose is not present n Forms a loop preventing transcription n The same protein has opposite affects depending on the environment

The Loop (negative control)

Transcription in Eukaryotes: n Many of the problems are similar to those in prokaryotes n However, often the actions of different tissues must be coordinated n Development and physiological changes

Cis acting sequences: Has multiple regions:

The core promoter: The RNA polymerase II binding site n Transcription start sequence n TATA box 30 bp up stream of initiation site n

Promoter-proximal cis acting sequences Helps binding of RNA polymerase II to the promoter n CCAAT box and GC rich segment n 100 -200 bp up stream n

Distance independent elements: n Act at some distance from the promoter n n Enhancers: Increase transcription rates Silencers: Decrease transcription rates Typically they are cis acting sequences affected by trans acting regulatory proteins n The distance can be 50 kb or more from the promoter! n Can be up stream or down stream from the promoter n

Mechanism of action from a distance: n How can they function so far away? n DNA forms loops n Proteins bound to regulatory regions are brought into contact with the promoter region

Action from a distance

Tissue specific transcription: n Some genes only turn on in particular tissues n Regulatory substances may only be present in certain tissues n Can be a repressor or an enhancer n The complex regulation requires a large number of regulatory genes n Hormones

Reporter genes and gene regulation: n n n Splice in a reporter gene A reporter gene produces an obvious phenotype Presence of the reporter substance indicates one or more enhancers must be near Moving the spliced bit localizes the enhancer We gain knowledge of where an enhancer works In time we can isolate trans acting regulatory compounds

Reporters

Regulatory mutations from chromosomal rearrangements: n Can be due to chromosomal rearrangements n Move the enhancer of one gene next to the transcription unit of another

Bar eye mutation n An eye regulatory sequence is placed next to a gene not normally expressed in the eye n This compound leads to the death of many cells in the eye

Tab (trans abdominal) mutation: n Genes normally only expressed in the abdomen are expressed in parts of the thorax