Prokaryotes
What does the lacZ gene encode for? What about lacY?
lacZ: encodes B-galactosidase
lacY: encodes galactoside permease
Multicellularity implies a need for _____ = different ____ have different functions even though they have the same ____.
Multicellularity implies a need for _differentiation__ = different __cells__ have different functions even though they have the same __DNA__.
What is the difference between promoter and enhancers/silencers?
Promoter: "region of DNA where RNA polymerase binds/begins transcription"
Enhancers/Silencers: "regulatory DNA elements bound by regulatory proteins"
-found upstream/downstream of promoter
From a single gene, RNA editing can result in...
1) different proteins in different cells
2) different proteins within the same cell
What is constitutive gene expression? How does it differ from regulated gene expression?
Constitutive: gene product is made CONTINUOUSLY
Regulated: gene produce made ON DEMAND; expression can be either induced OR repressed
How are LacZ and LacY transcribed?
As a single polycistronic mRNA
What is chromatin? What is a nucleosome? Why is chromatin packing important?
Chromatin: DNA binding proteins + DNA packaged together
Nucleosome: basic unit of chromatin packaging; composed of histone proteins (positively charged) + DNA
Important bc tight packing BLOCKS RNA polymerase access to DNA (prevents transcription)
What common features are present at all genes?
Basal transcription complex: composed of promoter sequence, RNA polymerase, & general transcription factors!
***very important***
_____ ______ produces more than one mature mRNA from the SAME gene.
_Alternative_ _splicing_ produces more than one mature mRNA from the SAME gene.
_____ is the preferred source of carbon for E. coli. However, if ____ is not available, bacteria can break down ____ to generate _______.
__Glucose__ is the preferred source of carbon for E. coli. However, if __glucose__ is not available, bacteria can break down __lactose__ to generate ____glucose___.
_____ induces expression of B-galactosidase.
_____ inhibits production of B-galactosidase.
_Lactose_ induces expression of B-galactosidase.
_Glucose_ inhibits production of B-galactosidase.
_____ must be opened up before a gene can be _____.
What are 3 ways to modify chromatin?
__Chromatin_ must be opened up before a gene can be __transcribed_.
1) DNA methylation
2) modification of histones
3) chromatin remodeling
What features vary from gene to gene?
Enhancers/silencers + the regulatory transcription factors they bind vary from gene to gene
-Enhancers/silencers can be located FAR away from the gene; upstream/downstream/within introns; 5' --> 3' orientation + exact location does NOT matter
1) RTFs bind to enhancer
2) recruitment of HATs & chromatin remodelers opens chromatin
3) assembly of general transcription factors @ promoter sequence
4) recruitment of RNA polymerase
5) transcription begins!
Fill in the blank:
_____ is a product of a ____ regulatory gene.
_____ binds the operator and blocks RNA polymerase.
_____ regulation of repressor.
Operon: "portion of DNA including a set of genes involved in a specific metabolic pathway"
Single RR: "contains operator + promoter"
_Repressor_ is a product of a __separate__ regulatory gene.
_Repressor_ binds the operator and blocks RNA polymerase.
_Allosteric_ regulation of repressor.
What is the difference between positive and negative regulation?
Positive: NO repressor normally on operator (still normally off); need activator protein to bind to promoter sequence to turn gene on
1) DNA methylation: DNA methyltransferases add methyl groups to cytosine --> condenses chromatin --> inhibits transcription
2) Modification of histones:
-ex: histone acetylation: HATs open chromatin (allow transcription to occur); HDACs remove acetyl groups (condense chromatin)
3) Chromatin Remodeling: use ATP to move nucleosome --> opens up chromatin --> allows RNA pol to access DNA (transcription can occur)
Tell me about regulatory TF's
Regulatory TF's have DNA binding domains, which can recruit co-regulators (HATs/HDACs)
-co-activator = HAT (opens chromatin)
-co-repressor = HDAC (condenses chromatin)
How does differential gene expression in cells occur?
Differential gene expression results fro production/activation of different regulatory proteins (RTFs)
1) when glucose levels HIGH, cAMP levels LOW
2) cAMP receptor protein (CRP) binds cAMP
3) cAMP-CRP complex acts as an activator of lac operon
-POSITIVE regulation! (high glucose = low cAMP = low levels transcription; low glucose = high cAMP = HIGH levels transcription)
What happens when lactose is present/not present?
-Lactose binds to LacI, which prevents repressor from binding to operator; RNA polymerase can bind to promoter and transcribe lac operon
Not present: LacI binds to operator and PREVENTS RNA polymerase from binding to promoter (no transcription)
Is DNA methylation inherited? Why/why not?
Essentially, yes. When methylated DNA is replicated, each strand becomes hemi-methylated (1 strand methylated), and then DNA methyltransferase recognizes this hemi-methylated DNA and methylates the other stand.
**methylation status of genes from previous cell generation is remembered**
HOWEVER, DNA methylations are "reset" during gametogenesis
How do enhancers get close to the promoter region of their intended gene if they are so far away?
CHROMATIN LOOPING: brings enhancers & promoters to close spatial proximity; *mediator complex* links distant enhancers to their promoter
Name & describe the types of post-translational forms of gene regulation
1) Molecular chaperones: assist proteins to fold properly
2) Proteasomes degrade upiquitin-tagged proteins:
-targets misfiled proteins = type of error correction mechanism
-controls protein lifespan = regulation of gene expression
3) chemical modification:
-ex: phosphorylation: proteins can be activated/deactivated by addition of phosphate group
4) protein transport: transport from cytoplasm to nucleus
-ex: STATs
When do high levels of transcription occur?
1) NEGATIVE control by Lac repressor (represses transcription unless lactose present)
2) POSITIVE control by CRP (activates transcription when glucose levels low)
High levels transcription seen when
1) repressor NOT bound to operator
AND
2) CRP bound to promoter (low glucose)