Gene Regulation Flashcards
Why do Bacteria need to regulate Genes?
- they are nutritional opportunists
- to do so, they’ve evolved regulatory systems that couple expression of gene products to sensory systems that detect relevant compound in enviro.
Basics of Prokaryotic Transcription
- Promoter & Operator
- Activators & Repressors
- Positive & negative regulation
-Promoter: where RNA polymerase binds - allows transcription to occur
-DNA seq near promoter serve as binding sites for seq-specific regulatory proteins called activators & repressors
-activators physically help tether RNA poly to promoter
-repressors either physically interefere w/ binding or impede movement of RNA polymerase
Operator: where repressors bind
Positive Regulation: when activator protein must bind to its target DNA site as necessary prerequisite for transcription
-the PRESENCE of bound protein needed for transcript
Negative Regulation: when repressor protein must be prevented from binding to its target site for transcript to begin
-the ABSENCE of bound repressor allows it to begin
Gene Regulation; Bacteria
-how bact. genes organised
- Regulatory gene
- role of regulatory protein
- have related gene together in clusters (OPERONS) so that genes required for similar functions can be made together
- Regulatory gene (may be close or far from operon) makes regulatory protein
- regulatory protein can either be inhibitory or enhancing to transcript
Gene regulation; Prokaryotes
- 2 ways regulation achieved
- Sigma factors
2 ways: Sigma factors or regulatory proteins
SIGMA FACTORS:
-recognizes genes by their promoters (recognizes specific set of sequences at -10 & -35 regions
-Therefore, set of genes required for particular function can have same promoter regions & be activated by a given sigma factor
Gene regulation; Prokaryotes
- Regulatory proteins (2 types)
- trans & cis acting factors/repressors
Regulation by REGULATORY PROTEINS
-regulatory proteins bind to target site - can control expression
Positive regulator: Protein binds & transcription starts (called positive control)
-trans-acting factors must bind to cis-acting sites for RNA polymerase to initiate transcript. at promoter
Negative regulator: Protein binds & transcription stopped/inhibited (or binding suppresses expression) (called negative control)
-trans-acting repressor binds to cis-acting operator to turn off transcription
Gene regulation; Prokaryotes
-Induction and repression
Induction: process where gene expressed in response to presence of specific molecule called an inducer
Repression: When the gene is switched off in response to presence of specific molecule called repressor
Gene regulation; Prokaryotes
-MalT operon - e.g. of positive regulator and induction
- regulatory protein ca only bind to DNA in specific conformation
- regulatory proteins change their conformation after binding w/ effector molecule (Maltose)
- regulatory proteins requires maltose to bind at regulatory site -> is a positive regulator (binding = expression)
- e.g. of induction because genes required for maltose catabolism normally silent if no maltose, but induced in presence of maltose
Gene regulation; Prokaryotes
-lac operon - e.g. of negative regulator and induction
- regulatory protein binds at regulatory site w/out effector molecule
- is an example of negative regulator -> binding or regulator stops gene expression
- example of induction because genes required for lactose catabolism induced only in presence of lactose (remain silent in absence of lactose)
Telling if something is a positive or negative regulator and whether it is induction or repression
- Induction/repression: look and see what the effect of a small molecule (effector) is
- presence of molecule induces transcript = induction
- presence of molecule turns off transcript = repression - Positive/negative regulation: see what regulatory protein does
- if naturally binds and stops transcript = neg
- if naturally binds and helps transcript = positive
The lac Operon - lacI as a negative regulator
-In presence of lactose and without
a) Lac operon w/out lactose
- LacI repressor binds at operator site & maintains lac operon in inactive condition (Operator lies between promoter & structure genes)
- prevents RNApoly from initiation transcription
b) Lac operon in presence of lactose
- Inducer binds to repressor and converts it to inactive form
- inactive repressor can’t bind to operator -> RNA poly allowed to move forward for transcription
The lac Operon - cAMP-CAP complex acts as positive regulator
-How it works as a positive regulator
- Catabolite activator protein (CAP) binds to promoter of lac oepron and stimulates transcription
- must bind w/ adenosine-3’, 5’-cyclic monophsophate (cAMP) before binding
- binding of cAMP-CAP to promoter activates transcription by facilitating binding of RNA polymerase
- lvls of cAMP inversely related to glucose - low glucose stimulates high cAMP (vice versa)
- ensures that cell uses glucose first, even if lactose present
Gene regulation by DNA bending - e.g. of CAP protein
- Active CAP recognizes specific seq of DNA for binding
- CAP binding induces DNA bending
- introduces sharp kink w/in binding sequence (results in 90 degree bend)
- CAP functions by interacting w/ RNA poly (alpha subunit)
- introduces sharp kink w/in binding sequence (results in 90 degree bend)
is very important to start transcription
Gene regulation by DNA looping - using ara operon as an e.g.
- Structure of ara operon
- In Absence of arabinose
- in presence of arabinose
- ara operon has 3 structural genes: araB, araA, araD
- regulatory gene = araC
- 4 regulatory regions: araO1,araO2, araI1 & araI2
a) Absence of arabinose
- AraC binds to I1 & O2 - causes DNA looping
- results in no transcription from BAD and C
b) Presence of arabinose - Arabinose interacts w/ AraC - changes affinity
- AraC binds at I2 and of CAP at it’s binding site is required for transcription
*AraC acts as both positive & negative regulator
Gene regulation by attenuating transcription - using trp operon as e.g.
- Structure of trp operon
- When tryptophan levels are low
- trp operon has 5 structural genes: trpE, D, C, B, A
- control region has promoter, operator, leader peptide coding region (L) & attenuator
- region L locate between operator and attenuator
- control region has promoter, operator, leader peptide coding region (L) & attenuator
- tryptophan is needed by the cell!
a) Tryptophan lvls low - RNA poly beings to transcribe DNA, producing region 1 of 5’ UTR
- ribosome attaches to 5’ end of 5’ UTR and translates region 1 while region 2 being transcribed
- ribosome stalls at trp codons because lvls low, region 2 is not covered by ribosome when region 3 transcribed
- when region 3 transcribed, paired w/ region 2. When region 4 transcribed, it cannot pair w/ region 3 (as already paired w/ region 2); attenuator never forms and transcription continues
Gene regulation by attenuating transcription - using trp operon as e.g.
-When tryptophan levels are high
b) Tryptophan lvls high
- RNA poly beings to transcribe DNA, producing region 1 of 5’ UTR
- ribosome attaches to 5’ end of 5’ UTR and translates region 1 while region 2 being transcribed
- RNA poly transcribes region 3. DOES NOT stall at trp codons, because it is abundant
- ribosome covers part of region 2, preventing it from binding w/ region 3. Region 4 transcribed and pairs w/ region 3, producing attenuator that terminates transcription
