Gene Regulation II

What were the four tools available to Jacob and Monod?

1. Mutants

2. Construction of merodiploids

3. Assay for enzyme activity

4. Logic

What are the two types of mutants?

1. Constitutive

2. Non-inducible

What did construction of merodiploids allow for?

Complementation

What could an assay for enzyme activity tell you?

If a gene was being expressed or not

What is a constitutive mutant?

It is where the system is always on whether or not lactose is present

What is non-inducible mutant?

Could not induce the operon (the system was never on) whether lactose was present or not.

What are merodiploids?

It is partially diploid on a small region of its chromosome.

What is an uninduced state? Describe what happens

It is when the system does not want to be turned on. LacI (repressor) will bind to the operator and no transcription will occur as it is a road block and no genes will get expressed.

What happens in a non-inducible mutant?

LacI(s) is a super repressor that is present which prevents the repressor (LacI) from binding to lactose. It will bind to the operator and represses the operon.

What happens in the induced state?

The inducer (lactose or IPTG) binds to LacI which prevents it from binding to the operator. There is transcription of the operon and genes are expressed

What happens if you delete or mutate LacI?

Then it cannot repress and expression of the lac operon will occur

What is the Oc?

It is a mutation in the operator sequence that prevents the repressor from binding to the operator whether or not lactose is present

What is a cis-acting locus? What is an example?

It is a genetic region affecting the activity of genes on that same DNA molecule. lac operator

What is a trans-acting locus? What is an example?

Encodes for a factor that can act elsewhere. LacI

Why is LacI a trans acting locus?

This is because it is made into a protein at a distance that eventually binds to the operator.

Would a mutation in LacI be cis acting locus or trans acting locus?

Trans acting locus

What is a complementation test used for?

To see if two genes of interest are placed in the same chromosome or not. Or if two mutations are in the same gene or not

If the two genes are complementing, what does this mean?

That the mutations are in different genes but if they are not complementing then they are in the same genes

What is complementation? What is the complementation test the basis of?

When you take away a gene. Blue white screening

What are activators? Are they positive or negative control?

They are proteins that bind specific sequences of DNA. They are positive control

Do activators make contact with RNA polymerase? What is their function?

Yes and they recruit holoenzyme to the promotor. They also aid and facilitate the process of formation of the open complex for gene expression

What are activators classified by?

Structure of DNA binding domain

Is there a slight bind in the DNA?

Yes there is

What part of the DNA do activators bind to? What is induced?

Two major grooves and induces a conformational change

What is a target?

The part that interacts with the DNA (major groove binding site)

What is causing conformational changes?

Hydrogen bonding and ionic interactions. The protein is doing work and impacting the structure.

What sugar do cells use first? Why? Does bacteria use a bunch of different carbon sources?

Glucose because it feeds into a lot of biosynthetic pathways such as glycolysis and yes

What does the presence of glucose do? What does glucose up-regulate?

It represses many genes. The activity of adenylate cyclase which makes cAMP

What does cAMP bind to?

Catabolite activator protein (CAP)

What does CAP do?

It is an activator that recruits RNA polymerase to promoters that would not be turned on efficiently otherwise

What does different types of genes being turned on based on?

Environmental context

What happens if no glucose is around?

Xylose is sensed and biosynthetic genes are turned on for xylose degradation