Lecture 7 Flashcards
What is a Gene?
The sum of all transcribed and non-transcribed regions that are necessary to properly express a gene
What is the Transcription unit?
The transcribed region of a gene
What is the Open Reading Frame (ORF) of the transcription unit?
The Coding Region, the part that has a start codon and the stop codon
What have more than one ORF?
Operon-derived mRNAs have more than one ORF
What are examples of RNA that are not translated?
rRNA, tRNA, microRNA and long non-coding RNAs
What are Untranslated Regions (UTR) and introns?
Parts of a gene that are transcribed but not necessarily coding
Where are regulatory sequences commonly located?
In the non-transcribed regions of a gene, but sequences with important regulatory functions may reside in the transcription unit and coding region
What does Transcription occur in?
The transcription bubble
What is Transcription carried out by?
RNA polymerase
What is the reason for the Transcription bubble?
Closing the DNA right away will prevent accidental transcription by other factors
What is the Non-template Strand of the DNA known as?
The Coding strand and the Sense strand. It is equivalent to the mRNA
What is the Template Strand of DNA known as?
The Noncoding strand and the antisense strand
What is the Key difference between Bacterial and Eukaryotic gene regulation?
The ground state of bacteria is on and the ground state of eukaryotes is off
Why is the Ground state of genes in Eukaryotic organisms off?
Because they are tightly wound around nucleosomes and are inaccessible from DNA binding proteins
Why is the Ground state of genes in Bacteria on?
Because the same nucleosomes and histones are not present in bacteria as they are in Eukaryotes so the DNA is naked and accessible
How do Bacteria turn off a gene?
They must actively deploy a repressor protein
How many gene products are needed for the core enzyme of bacterial RNA polymerase?
4
What are the 5 parts of Bacterial RNA polymerase?
2 alpha subunits, omega, beta and prime
What are the two alpha subunits encoded by for Bacterial RNA polymerase?
The same gene
What is the Sigma actor needed for?
Specific binding of the RNA polymerase to the promoter
What is Holoenzyme?
The Core enzyme of bacteria RNA polymerase plus Sigma factor
Where are Promoters found?
In eukaryotes and prokaryotes
What is the Promoter?
A region of a gene containing regulatory sequences that are close to (or contain) the transcription start site
What is the Promoter required for?
Transcription initiation via RNA Polymerase
What does the Promoter region have?
Binding sites for the RNA Polymerase and helper factors like sigma factor
What is the function of the terminator region?
It terminates bacterial transcription
Which organisms is the Terminator region found?
In bacteria only
What occurs in Positive Regulation?
An activator protein involved that recruits RNA polymerase
What occurs in negative regulation?
An Repressor protein that binds to the operator sequence and interferes with transcription by blocking RNA polymerase
What is Basal Transcription?
When RNA polymerase hasn’t been recruited specifically but there is no repressor to stop it. It is unregulated
What are Ligands?
Small molecules that bind to activators and change the confirmation of the protein which promote the binding of DNA
What are Activators and Repressors regulated by?
Ligands (hormones, small peptides or metabolites etc)
What is an Allosteric site?
A region in a 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
How is Glucose formed in E. coli?
Lactose is brought into the cell the by b-Galactosidase and then breaks the lactose into allolactose or then to Galactose + Glucose
How many Genes is the Lac operon composed of?
3
What are the Genes of the lac operon?
LacZ
LacY
LacA
What does the lacI encode for?
The repressor protein which is the regulatory portion
What does the lacZ encode for?
B-galactosidase
What does the LacY encode for?
Permease that transports lactose into the cell
What is an Operon?
A group of genes that are adjacent to each other and transcribed together because they are controlled by a single regulatory region
What is the product of the operon?
A polycistronic mRNA
What is polycistronic mRNA?
An mRNA that encodes multiple proteins
What is an Operator?
A sequence of DNA that is the binding site for a repressor protein which prevents transcription by preventing RNA polymerase from binding or blocking it
Where are operators found?
In bacteria
What does CAP stand for?
Catabolite activator protein
What is the Lac Operon regulated by?
Ligands b-1,6-allolactose (which blocks the activity of the lac repressor) and cAMP activates CAP
What conditions must be met for the lac operon to be induced?
Lactose is present and glucose is absent
What occurs in the Lac operon when glucose and Lactose are both present?
Basal Transcription
What occurs when glucose is present and lactose is absent?
The operon is repressed
What occurs when glucose is absent and lactose is absent?
The operon is repressed
What is Basal Transcription?
When RNA polymerase binds to DNA in the absence of activators and repressors
How does the lac repressor work?
A homotetramer and binds to two lac operators at the same time
Why is there no Transcription in the absence of Lactose?
Because allolactose is a product of B-galactosidase working on Lactose and allolactose is a ligand of the repressor protein which causes it to be inactive
Why does the lac operon not work in the presence of glucose?
Glucose inhibits the synthesis of cAMP by inhibiting the enzyme that produces it
What is cAMP?
A ligand that activates the CAP protein which recruits the RNA polymerase
How does cAMP work?
It changes the shape of CAP by rotating the DNA-recognition helices
What is CAP?
An activator that binds to the CAP site of the lac promoter
Why can basal transcription occur when there is a low presence of glucose?
Because the repressor protein is only bound to the operator 96% of the time which gives opportunity for basal transcription to occur
What was the approach by Jacob and Monod in figuring out the function of the different portion of the Lac Operon?
They used a plasmid called F-factor to make a partially diploid E. coli since E. coli is haploid. Then they cause mutations of these parts and saw the outcome
What does the Cis-Regulatory element refer to?
A small section of DNA important for regulation of a gene. Cis indicated that is is located on the same piece of DNA as the gene that it regulates, usually in the vicinity of the gene
What does the Trans-acting factor refer to?
A regulatory protein that binds to cis-regulatory elements or contributes to the regulation of a gene is some other fashion. The gene encoding the trans-acting factor does not need to be on the same piece of DNA
What are Trans Acting factors of the Lac Operon
The proteins of Cap and the Lac Repressor
What are the Cis-regulatory elements of the Lac Operon
Cap binding site and the Operator
What is Constitutive activity?
A situation where a gene/enzyme is continuously active regardless of the biological condition. Even it is on it should be off
What are the Structural genes of the Lac Operon?
LacZ, LacY, LacA
Why are LacZ, LacY, LacA structural genes?
Because they do not directly affect the regulation of the operon
What are the Regulatory elements of the Lac Operon?
Lac Repressor, lac operator, CAP site, promoter elements
What are Inducers?
Small ligands that bind to proteins and increase gene activity
What are the Inducers for the Lac Operon?
Allolactose for the lac repressor and cAMP for the CAP
What is the opposite of an inducer?
Co-repressor which are small ligands promoting repression
Where is the term inducer used?
Only in bacteria
What happens when B-galactosidase digests X-gal?
It turns blue
What occurs in the lacI-, type I mutation?
No lac repressor is made
What occurs in the lacI-, type III mutation?
Lac repressor is made but is not capable of binding to the operator (so the DNA binding domain is effected)
What occurs in the lacIs mutation?
The Lac repressor is made but cannot bind to allolactose or IPTG (mutation affecting allosteric site)
What is the acting of the Operon?
Cis-Regulatory
What is the acting of the Lac repressor?
It is Trans acting
What does the Is mutation result in?
A non inducible operon because Is is dominant to I+
What kind of Morph is the Is mutation?
An anti morph
