Genetics chp. 12 Gene regulation

Structural genes

encoding proteins

Regulatory genes

encoding product that interacts with other sequences to affect transcription and translation of those genes

Regulatory elements

DNA sequence that are not transcribed but play a role in regulating other sequences

Levels of Gene expression

1) Alteration of structure

2) Transcription

3) mRNA processing

4) regulation of mRNA stability

5) Translation

6) Posttranslational modification affecting protein stability

Expression types

• Constitutive expression: Continuously expressed under normal cell conditions

• Positive control: Regulator protein stimulates gene expression

• Negative control: Regulator protein inhibits gene expression

Operon structure

• Promoter: Upstream of structural genes and controls their transcription

• Structural genes: series of genes downstream of promoter that are transcribed into mRNA and then made into enzymes that will carry out biochemical reactions

• Operator: between promoter and structural genes and is a binding site for the regulator protein

Regulator sequence

Gene sequence controlling expression of structural genes by increasing or decreasing their transcription. Has its own promoter that is not apart of the operon

Regulator protein

Regulator sequence has its own promoter that will eventually enable synthesis of regulator protein that binds to an operator.

Operator

Genetic sequence that overlaps the 3’ end of promoter and 5’ end of structural genes. Its a binding site for the regulator protein that will increase or decrease transcription of structural genes.

Inducible and Repressible operons

• Inducible operon: Transcription is usually off and needs to be turned on (induced)

• Repressible operon: Transcription is usually on and needs to be turned off (repressed)

Negative inducible operon

Operon is negatively controlled and inducible so something is inhibiting gene expression but transcription needs to be turned on.

Negative inducible operon process

A regulator protein binds to the operator which overlaps with the promoter causing RNA polymerase to be blocked and unable to bind, preventing transcription. An inducer must be present for transcription to take place.

Inducer

Small molecule that turns on transcription

Precursor V

Small molecule acts as an inducer by binding to the regulator protein and altering its shape to not allow it to bind to the operon. When this happens, RNA polymerase can bind to the promoter and transcription can take place.

Negative repressible operon

Control of the operon site is negative but the operon is usually on and must be turned off.

Corepressor

Small molecule that binds to the repressor to make it capable of binding to the operator site that inhibits transcription

Negative repressible operon process

The regulator protein acts as a repressor that will inhibit transcription of operon but cannot bind to the operator site on it’s own. Transcription will occur at first and product U will be in a abundance acting as a core repressor that will allow the repressor to bind and stop transcription therefore not allowing for structural genes to be synthesized into proteins. When U runs out transcription will start again and the cycle continues.

Positive inducible operon

Control of the operator site is positive but transcription is usually off and needs to be turned on.

Positive inducible operon process

A regulator protein acts as an inactive activator that cannot bind to the promoter on its own so a small molecule binds and allows for the activator to bind to the promoter site and turn on and increase rate of RNA polymerase activity.

Positive repressible transcription

Control of the operator site is positive but the operons transcription is usually on and needs to be turned off.

Positive repressible transcription process

The regulator protein is an active activator that allows for transcription to be turned on at a high rate. The products produced in the biochemical pathways bind to the activator and change its shape to not allow it to bind to the promoter therefore turning transcription off

inducer for lac operon of E. Coli

allolactose

E coli. repressor encoding gene

Lacl

E. coli operon promoter sequence

lacP

E. coli operon operator sequence

lacO

E. coli structural genes

lacZ: encodes for B-galactosidases

lacY: encodes for permease

lacA: encodes for transacetylase

Lactose metabolism process

Lactose does not easily diffuse across the bacterial membrane so lacY encodes for permease which transfers lactose into cell. B-galatosidase produced by the lacZ gene then converts lactose into galactose and glucose. B-galactosidase also converts lactose into allolactose which acts as an inducer in lactose metabolism.

classifying the lac operon

Negative inducible operon. Regulator protein is an active repressor that binds to the operator (lacO) and stops transcription.

Process of lac operon is presence of lactose

Lactose is turned into allolactose by B-galactosidase which acts as an inducer that binds to the repressor. The repressors shape changes and cannot bind to the lacO (operator) sequence therefore allowing RNA polymerase to bind to the promoter and let structural genes be transcribed to eventually become enzymes.

What type of bacteria was used to determine lac operon function and structure

Merezygotes/partial diploids: Full bacterial chromosome and an Fplasmid containing extra DNA

Cis vs trans mutations to lac operon

• Cis-acting mutation: Able to control the expression of genes when on the same piece of DNA ex. promoter or operator

• Trans-acting mutation: able to control expression of genes on other DNA molecules ex. regulatory gene

Structural gene mutation

affect structure of enzyme but does not affect the regulation of their synthesis

Regulator gene mutations (trans acting)

affects the regulation of protein production

different lac gene affcects

• lacl- leads to constitutive transcription of structural genes

• lacl+: dominant over lacl- and is trans acting. Normal regulation of lac operon is brought on when lacl+ is present

• lacls: leads to superrepressor mutation and is most dominant so transcription is prevented

Operator mutations

lacOc: constituve

Promoter mutations

lacP-: cis acting

summary of all genes

• lacP: promoter sequence

• lacO: operator sequence

• lacZ: sequence encoding B-galactosidase

• lacY: sequence encoding permease

• lacA: encodes for transacetylase

• lacl- leads to constitutive transcription of structural genes

• lacl+: dominant over lacl- and is trans acting. Normal regulation of lac operon is brought on when lacl+ is present

• lacls: leads to superrepressor mutation and is most dominant so transcription is prevented

• lacOc: constitutive transcription

Catabolite repression

use of glucose when available and repressing the metabolite of other sugars

is catabolite repression a form of negative or positive regulation

Positive regulation

cAMP

concentration of cAMP is inversely proportional to the level of available glucose.

More cAMP means lower lvls of glucose and the inverse

Trp operon

controls synthesis of amino acid tryptophan

Trp operon classified as

Negative repressible operon

Trp Operon process when tryptophan levels are low

the regulator protein is an inactive repressor so transcription of five structural genes takes place.

Trp Operon process when tryptophan levels are high

Tryptophan binds to the inactive repressor which activates it and allows it to bind to the operator stopping RNA polymerase from transcribing the five structural genes and therefore lowering the levels of tryptophan

Requirements for minimal levels of transcription

• General transcription factors

• RNA polymerase

Requirements for normal levels of transcription

• General transcription factors

• RNA polymerase

• Transcription regulator proteins

Transcription regulator proteins

activators or repressors that can bind to enhancers or silencers

Activator proteins

• Stimulate and stabilize basal transcription apparatus by binding regulator sequences like enhancers

Insulator

DNA sequence that blocks effects of enhancer

Repressors

Bind regulatory sequences on promoter to inhibit transcription

Two parts of promoter sequence

• Activator binding site: Transcriptional activators (Activator or Repressor) bind

• Core promoter: Basal transcription apparatus binds

insulator process

If insulator is between a promoter and an enhancer than the action of the enhancer is blocked.

If the insulator is outside the region between the promoter and enhancer than no effect

Stressor process

Stressor activates transcription factors that bind to short DNA consensus sequences that will produce different proteins for stress response

Result of altering chromatin structure

Gene expression will be altered

Mechanisms for altering chromatin structure

• Changes in patterns of DNA methylation

• modifying histone proteins

• RNA molecules that affect chromatin structure and gene expression

Possible histone modification additions

• Phosphates

• Methyl groups

• acetyl groups

• ubiquitin