17: Gene Regulation

metabolic regulation

1. regulation of enzymatic activity (i.e., affinity, activation, inhibition)

2. regulation of enzyme synthesis (i.e., gene expression)

operons

unit of genetic function found in bacteria with 3 elements

• promoter

• operator (within the promoter region)

  • controls the access of RNA pol to genes

• cluster of genes (whose products function in a common pathway)

repressor when active

represses gene expression

• repressor binds to operator, blocking RNA pol from binding

• transcription decreases

elements of trp operon

• 3 reactions

• 3 enzymes

• 5 genes

• 1 promoter

• 1 operator

• 1 long mRNA

• 5 start codons

• 5 stop codons

  • = 5 ORFs

• 5 polypeptides in single mRNA

trp repressible operon

under normal conditions:

• operon is ON

• trpR is inactive

  • activated by concentration of tryptophan that activates trpR

corepressor

tryptophan

• cooperates with trpR (repressor) to turn off operon

anabolic pathway

operon: ON

• tryptophan expressed

increased [tryptophan]: binds to trpR and activates it

trpR binds to operator (within promoter)

operon: OFF

• tryptophan not expressed (already high concentration in the cell)

negative regulation

• trp operon

• lac operon

positive regulation

• lac operon (in the presence of glucose and lactose)

lac inducible operon

normal conditions

• operon is OFF

• lac repressor is active

  • inactivated as it binds to inducer, allolactose

elements of lac operon

• 1 promoter

• 1 operator

• 1 regulatory gene

  • lacI codes repressor

• 3 genes

  • lacZ: b-galactosidase

  • lacY: permease

  • lacA: transacetylase

catabolic pathway

lac operon OFF

lac repressor is active

• lactose present

• inducer allolactose binds to lac repressor to inactivate it

lac repressor inactive

lac operon ON

• transcription by RNA pol II

• increased lactose pathway enzymes (b-galactosidase)

• hydrolysis of lactose

lactose → glucose + galactose

lac operon: positive regulation

• cAMP binds to CRP (cAMP receptor protein)

• once activated, CRP activator binds to lac promoter

• recruits RNA pol

• CRP increases gene transcription of lac operon

+ glucose

+ lactose

• decreased AC

• decreased cAMP

• CRP inactive

• decreased affinity for RNA to bind to promoter

  • because glucose inhibits AC; no CRP

• operon OFF

• *some transcription is still possible due to presence of lactose

+ glucose

- lactose

• operon OFF

• lac repressor active

- glucose

- lactose

• operon OFF

• AC active

• cAMP activates CRP

• lac repressor active

- glucose

+ lactose

• operon ON

• AC active

• cAMP activates CRP

• increases affinity for RNA pol

• transcription of lac mRNA

eukaryotes

gene expression can be modulated at different stages

• does not involve operons

• depends on control elements

differential gene expression

cells from different cell types (tissues) have the same genome but a different program of gene expression

• depends on the cell’s specific

  • function

  • localization

  • differentiation

  • signals

  • internal and external environment

control elements

serve as binding sites for transcription factors and help recruit the transcription initiation complex

cis-acting DNA control elements

control elements located on the same chromosome as the gene they regulate

• promoters

• enhancers

trans-acting DNA control elements

control elements that bind to the gene they regulate

• transcription factors

enhancer

group of distal control elements that increase gene transcription

• upstream from promoter

• cis-acting control elements

proximal control elements

close to promoter

• upstream from the gene

• bind TFs to help gene expression

general TFs

can bind to

• promoter of all protein-coding genes (TATA box)

• other TFs

• RNA pol II

specific TFs

can bind to

• control elements (upstream and downstream)

• other TFs

• mediator proteins

which increase or decrease the rate of transcription

transcription initiation complex (TIC)

recruited by mediator proteins

• includes

  • RNA pol II

  • general TFs

  • other coactivators

steps in TIC complex

• TFs bind to control elements

• once bound, TFs interact with mediator proteins that act like bridges

• mediator proteins stabilize and recruit TIC

• DNA looping brings distal elements like enhancers close to promoter region so their bound proteins can interact with transcription machinery

coordinated gene expression

for enzymes that belong to the same metabolic pathway

• achieved by binding the same TFs to the same combination of control elements

differential gene expression example

• different TFs are active in different cells

• liver and lens cells

• both contain entire genome

  • albumin gene - important for liver cells

  • crystallin gene - important for lens cells

  • control elements required for both genes

• liver cells produce liver specific TFs that bind to albumin gene enhancers

• lens cells produce lens-specific TFs to do the same with crystallin

RNA interference

process in which small RNA molecules regulate gene expression by targeting specific mRNA molecules

1. degradation

2. translation inhibition

siRNA

complementary to mRNA target

• lead to degradation

• ex. dicer

miRNA

partially complementary to mRNA target

• forms hairpin structure structure processed into mature miRNA

• leads to translation inhibition

• ex. drosha

RISC (RNA-induced silencing complex)

1. siRNA or miRNA loaded into RISC complex

2. RISC uses RNA strand (siRNA or miRNA) as guide to find complementary mRNA

3. once it finds target:

   a. if siRNA - RISC cleaves mRNA =degradation

   b. if miRNA - RISC blocks translation or destabilizes mRNA