Lecture 7 Transcription and Gene Regulation Flashcards
What is a gene?
The sum of all transcribed and non-transcribed regions that are necessary to properly express a gene (must consider not just the coding region, but also the regulatory region etc)
The transcribed region of a gene is referred to as the:
Transcription unit
What is the open reading frame (ORF) of the transcription unit?
The coding region (operon-derived mRNAs harbor more than one ORF)
Some genes encode _____ that are no translated
RNAs
Not everything that is transcrbed is necessarily coding: provide an examp
5’ and 3’ UTR (untranslated regions) and introns
Regulatory sequences are commonly located in the _________ regions of a gene but sequences with important ______ functions may reside in the transcription unit and coding region
Regulatory sequences are commonly located in theNON-TRANSCRIBED regions of a gene but sequences with important REGULATORY functions may reside in the transcription unit and coding region
Transcription occurs via _____ _______
RNA polymerase
During transcription _____ the DNA is melted; Transcription then occurs in a “______”
During transcription INITIATION the DNA is melted; Transcription then occurs in a “TRANSCRIPTION BUBBLE”
What is the nontemplate strand?
Coding strand = “sense” strand Identical in sequence to the mRNA 5’-3’
What is the template strand?
Non-coding strand = “antisense” strand 3’-5’
Principle difference between bacterial and eukaryotic gene regulation?
- The ground state is ON in bacterial gene regulation (DNA is accessible at all times)
- Gene requires repression to turn “off”
- Whereas ground state in eukaryotes is OFF (because of nucleosomes = DNA is inaccessible)
- Gene requires activation “on”
Bacterial RNA Polymerase is a multi-subunit protein complex
- ____different gene products required for the ____ enzyme
- Two ___ subunits which are identical and encoded by the same gene
- ___, ____ and _____ are encoded by three additional genes
- _____ _____ is needed for specific binding of the RNA polymerase to the promotor
Bacterial RNA Polymerase is a multi-subunit protein complex
- Four different gene products required for the Core enzyme
- Two Alpha subunits which are identical and encoded by the same gene
- omega, beta and beta prime are encoded by three additional genes
- sigma factor is needed for specific binding of the RNA polymerase to the promotor
What is Holoenzyme?
Core enzyme (of RNA polymerase) plus Sigma factor
Sigma factor has the ability to bind to the ____ ____ _____ which drastically improves the ability to bind to promotors
Sigma factor has the ability to bind to the Core RNA Polymerase which drastically improves the ability to bind to promotors
What is a promoter?
Region of a gene containing regulatory sequences that are close to (or contain) the transcription start site
Required for transcription initiation via RNA polymerase
What is positive regulation?
Involves an activator
- activator bind to activator binding site
- allows RNA polymerase to bind to promotor
What is negative regulation?
Involves a Repressor
- Bind to operator sequences (part of promoter)
- Either prevents binding of RNA polymerase OR blocks progress of RNA polymerase
What is the Operator Sequence?
- part of promoter
- designed to interfere with transcription (ie turns gene off)
What is a ligand? What is it’s role in transcription?
Small units (metabolites/hormones/peptide hormones) that can bind to activators (via ligand-binding site/allosteric site) = change conformation of protein = Activating: Promote binding of DNA or Inhibiting: Promote release of DNA from activator-binding site
What is an allosteric site?
Region of protein to which a small molecule can bind (the ligand) resulting in a conformational change of the protein that modifies the activity of the protein
The result can be inhibitory or activating
The lac operon comprises three genes:
Beta-galactosidase
Permease
Transacetylase
What are operons?
A group of genes that are adjacent to each other and transcribed together (under control of a single regulatory region)
What result on mRNA does an operon have?
Resulting mRNA is polycistronic
Are operons found in eukaryotes or bacteria?
bacteria
Very rare in eukaryotes
An mRNA that can encode more than one polypeptide separately within the same RNA molecule is called:
Polycistronic
What are operators?
Are they found in bacteria or eukarya?
Region of DNA that acts as a binding site for a repressor protein
- Found in bacteria (has not been identified in eukaryotes)
What are the three structural genes of the Lac operon? What do they do?
- lacZ: beta-galactosidase
- cleaves lactose into galactose and glucose
- also produces beta1,6-allolactose
- lacY: transports lactose into the cell
- lacA: transfers acetyl group to certain sugars
What are the two regulatory genes of the Lac operon?
- lacI: lac repressor - not part of the lac operon but the lacI gene lies nearby (upstream of lac operon)
- CAP: catabolite activator protein
- also known as CRP (cAMP receptor protein)
What are the two Substrates/Effectors (ligands/inducers) of the lac operon?
- beta-1,6-allolactose: a transient byproduct of beta-galactosidase activity
- Triggers conformational change in lac repressor
- cAMP: cyclic adenosine monophosphate
- Allosteric effector for CAP/CRP - made when glucose is low
What two conditions must be met to have the lac operon induced?
- Lactose is present in the growth medium
- Glucose is absent from the growth medium
If glucose is present in the growth medium what happens?
Glucose is preferred sugar: Basal transcription
What is Basal transcription?
- occurs when RNA polymerase binds to DNA in the absence of activators and repressors (RNA polymerase Is neither specifically recruited to nor prevented from binding to the promotor)
What is the general mechanism of operator function?
- competition for promotor occupancy
- operator-bound repressor blocks binding or transcription of RNA polymerase
- Operator doesn’t have to be located between -10 and -35 elements
- Basal level of transcription generally refers to random binding of RNA Pol to the promoter in the absence of activators and repressors
Under what conditions of the lac operon might you see basal transcription?
Basal transcription can occur in a repressed state
Lac operon overview:
If we have neither the CAP(activator) nor Lac repressor what do we see?
Can happen in ___glucose, ____lactose conditions
Basal transcription - RNA polymerase will bind occasionally
Can happen in _+_glucose, _+_lactose conditions
In +glucose, -Lactose conditions what would we see?
No lactose in the medium = attracts binding of Lac repressor = operon is off
What would happen in -glucose, -lactose conditions?
No lactose = attracts Lac repressor = Operon off
What would we observe in -Glucose, +lactose conditions?
Operon ON = this is the only state where we see a fully activated operon
Presence of lactose in the absence of glucose = CAP binds and repressor is removed
