Gene Regulation Flashcards
Utility of regulating gene expression
- Differentiate gene expression allows cells to carry out specialized functions
-Prok
Allows cells to rapidly adapt to changing env
Energetically economical (do not waste energy making genes that are useless in a particular env)
-Euk
Have specified tissues with various functions
Allows the tissue to express different genes from the same manual
Transcriptional regulation in Pro
- Proteins that bind to promoter
Activator -protein involved in activation of transcription
Co-activator
Repressor-protein involved in repressing transcription
Co-repressor
-Consequence to transcription
To induce -enhances tx of RNA
To repress -inhibits tx of RNA
These terms refer to what is happening to the expression of the RNA
Positive regulation
- Transcription is accelerated
- Requires activators
- INDUCIBLE activator (activator is activated - by co-activator; results in induced transcription
REPRESSIBLE activator(activator is repressed by inhibitor, transcription is repressed)
Negative regulation
- Goal is to inhibit tx
- Requires repressors
- Gene expression is induced (turned on when repressor is REMOVED from DNA)
- Gene expression is repressed (turned off when repressor binds to DNA -due to co-repressor activating the repressor)
Repressors and Activators - Four Pathways
Negative - REPRESSOR
INDUCIBLE (inactivation of repressor is required) Tx is on
REPRESSIBLE (activation of repressor required)Tx is off
Postive - ACTIVATOR
INDUCIBLE (activation of activator required)Tx is on
REPRESSOR (inactivation of activator required) Tx is off
What is an operon?
Multiple genes clustered and subject to regulation by a single promoter.
Operator
Region of DNA to which repressor can bind
Promoter
Region of DNA upstream of genes that recruit tx machinery
Lac Operon
- Three genes under regulation of single promoter
- Genes are all needed from the metabolism of lactose (a sugar)
E. coli prefers to exhaust it’s glucose reserved first before metabolizing other sugar
Goal is to only transcribe Lac genes when: absence of glucose and presence of lactose
*Need to induce tx when lactose is present
Presumably glucose has been depleted
Structure of lac operon
- Utility of arranging genes into an operon as a single promoter can regulate the expression of all the genes at once
- the promoter is “activated” all gene tx is enhanced
- If promoter is “repressed” all the gene tx is off
Lac operon is mRNA is polycistronic
z- Beta galactosidase - cleavage of lactose
y- permease - entry of lactose into cell
a -transacetylase - unknown function; necessary for lactose breakdown
Lac Z
Allolactase is byproduct of breakdown with lac Z
Lac I gene
- Produces the repressor of lac operon
- Placl - LacI promoter; produces lacI constitutively (all the time)
- Does NOT share tx regulation with Lac operon genes (independent)
Negative control
Transcription - OFF initiation is inhibited
Function of Lac I (ex. inducible repression)
- Lac I will bind to the Lac operator when there is NO lactose present in environment
- If there is no lactose, it doesn’t make any sense top produce proteins that function to break down lactose
Lac I (repressor active)
- When lactose is absent from env or very low concentration the repressor is active BUT… LacI binding is not permanent, so the cell will do “leaky/trace” tx ….minimal tx
LacI block tx
lac I protein binds to lac O DNA sequence; inhibits RNA Polymerase from binding
Reversal of NC
Reversal of NC - high lactose conditions
Inducer inactivated the repressor
TX is permitted -induction of lac operon
Positive control
Tx ON initiation is accelerated
Low glucose conditions
-Need to accelerate genes for metabolizing lactose
If glucose GONE/LOW, cell must shift to using other sugars (like lactose) exclusively for survival -cell will induce expression of Lac genes through use of inducible ACTIVATOR
PC- induced activator stabilizes
RNA polymerase binding at promoter
No PC
- High glucose conditions
- Activator remains inactive; tx is not enhanced
cAMP production is low, activator is inactive
what is glucose and lactose are both present in moderate amounts?
- If they are both present, moderate tx of Lac genes occurs because lacI is not bound to DNA, but neither is cap
- If there is a condition where LacI is bound to operator, and there is an absence of glucose -the lacI repressor trumps the activity of CAP
Types of Lac Operon Mutations
I- - LacI repressor is mutated so it cannot bind to operator
Is - LacI supersuppresor; cannot be removed from DNA by inducer
Oc 0 Operator is mutated so that the LacL repressor cannot bind
P- -Promoter sequence mutated; RNA polymerase cannot bind the promoter
CAP- -Cap protein is mutated so it cannot bind theDNA
Z -, Y-. A- - Genes for lactose metabolism are not functional
Cis vs Trans Mutants
- A mutation is “cis” in the DNA sequence is mutated which inhibits protein—DNA interaction (only affects the operon with the cis mutation)
-Oc Operator is mutated so that the LacI repressor cannot bind
P- Promoter sequence is mutated; RNA polymerase cannot bind the promoter - A mutation is “trans” is the protein is mutated; protein function affected (affects any operon in the cell)
I- LacI repressor is mutated so it cannot bind to operator
Is LacI superrepressor; cannot be removed from DNA by inducer
CAP- Cap protein is mutated so it cannot bind the DNA
Z-, Y-, A- Genes for lactose metabolism are not functional
Partial Diploids
BOTH genomes produce functional LacI
Top operon has functional operator; in presence of lactose, repressor removed from DNA
Moderate transcription
Produces mutant LacZ enzyme
Bottom, operon has mutant operator; LacI can never bind—NEVER repressed
Moderate transcription always
Produces functional LacZ—makes inducer of LacI
Ex: lacI lacO lacZ/lacl LacOLacZ
Euk regulation of gene expression
Eukaryotes do not produce polycistronic mRNA
One geneone protein
One promoter one mRNA
Organize coordinated gene expression through homology of promotor elements for several different genes
Coordinated genes all have same regulatory elements
Genes are independently transcribed (different locations throughout the genome)
Euk
- cis-acting DNA elements (DNA binding sites)
- trans-acting factors/proteins (transcription factors, repressors or activators)
- Enhancers – binding sites for activators (example: UASg, HREs – Hormone Response Elements)
- Silencers – binding sites for repressors (example: some HREs – Hormone Response Elements)
GAL genes
- Although each gene has its own promoter, each promoter has a similar structure (UAS elements); all three genes are expressed under the same cellular conditions
- GAL 7, GAL10, GAL1 -> metabolism of galactose in eukaryotes
- Upstream activation sequences elements are locations where enhancers bind
No galactose present
- GAL80 represses activity of GAL4 gene
- GAL3+galactose removes OR changes shape of GAL80 = GAL4 activation of tx
Regulation by Gal4 and Gal 80
Enhances action at the core promoter
Hormones can be inducers or co-repressors for coordinated gene expression
- Steroids
- ADH or Vasopressin
- Oxytocin
- Peptides (insulin)
Lac operon regulatory genes
CAP and cAMP
Gal genes regulatory genes
Gal4 activator
Gal3 + galactose (co-activators)
Lac O
in charge of transporting lactose into the cytosol and digesting it into glucose
