Lecture 3-Gene Expression and Transcription

Question 1

What does transcription produce? What is a transcription unit?

Answer 1

• An RNA strand complementary to a DNA strand template.
• A transcription unit is a portion of DNA that encodes for an RNA molecule and the sequences necessary for its transcription.

Question 2

What does RNA polymerase add to create mRNA during transcription? In which direction are they added? Which way does the RNA polymerase read the DNA template strand?

Answer 2

• Nucleoside triphosphates are added from the 5’ end to the 3’ end of the mRNA strand.
• RNA polymerase reads the DNA from in the 3’ to 5’ direction.

Question 3

What are the three steps in the transcription cycle?

Answer 3

1. ) Initiation
2. ) Elongation
3. ) Termination

Question 4

Where does RNA polymerase know where to bind to the DNA for transcription? Where are they located?

Answer 4

• RNA polymerase bind to promoter sequences located upstream from the genes.

Question 5

What is the symbol for the transcription start site? What is the downstream signal and what is the upstream signal?

Answer 5

• (+1) is the transcription start site.
• Downstream = (-)
• Upstream = (+)
• There is no zero.

Question 6

What are sigma factors? What do they do? Are they found in both prokaryotes and eukaryotes?

Answer 6

• Sigma factors help RNA polymerase bind to the promoter region in prokaryotes.
• Only found in prokaryotes.

Question 7

Is there more than one type of sigma factor? If so, why?

Answer 7

• There are multiple sigma factors because different sigma factors help RNA polymerase bind to different genes depending on the outside environmental signals.
• Ex.) Sigma factors for stress response, metabolism, general, etc.

Question 8

Where are the 2 consensus sequences located for sigma factor 70?

Answer 8

• There is a consensus sequence downstream from the promoter at -35 and one at -10.

Question 9

Do prokaryotes have multiple RNA polymerases to transcribe all of their genes? What is another name for RNA polymerase in prokaryotes?

Answer 9

• No. They only have one type of RNA polymerase.

- RNA polymerase is also known as the core enzyme without its sigma factor.

Question 10

What happens during prokaryotic transcription elongation?

Answer 10

• The sigma factor disassociates from the DNA template strand as the RNA polymerase moves down the strand creating the mRNA.
• The DNA strand is being unwound in front of the RNA polymerase but also being re-wound behind it after being transcribed.

Question 11

What is rho-independent termination?

Answer 11

• When the core enzyme is coming to the end transcription, there is a group of repeated G-C rich regions which then the mRNA folds back on itself (inverted complementary strands) forming a hairpin loop.
• This stalls the RNA polymerase as it begins to transcribe an A-T rich region. This complementary U-A region weakly interacts.
• The weak interaction between the U-A region and the stalling of the RNA polymerase is unstable enough to disassociate the core enzyme from the DNA strand, creating the mRNA strand.
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Question 12

What is an operon? What does polycistronic mean?

Answer 12

• A group of genes that are regulated by a single promoter and form into a single mRNA molecule.
• Polycistronic is a mRNA molecule that encodes for several proteins. Usually found in prokaryotes.

Question 13

What is the trp operon? What happens when the concentrations are high or low?

Answer 13

• The trp operon is an operon with 5 structural genes which encode for the amino acid tryptophan.
• It is always on unless tryptophan levels become too high. Tryptophan acts as the corepressor. A regulatory gene produces an inactive repressor and tryptophan binds to this repressor to activate it. The repressor binds to the operator site and blocks RNA polymerase from binding, not allowing transcription to occur.
• When tryptophan levels are low the inactive repressor cannot bind to the operator and transcription is not blocked.

Question 14

Do Eukaryotes use many different RNA polymerases for transcription? If so, what is the main one?

Answer 14

• Yes. Many different RNA polymerases are used.

- The main one is RNA polymerase II.

Question 15

What does in vitro mean and what is required for RNA polymerase to transcribe in vitro?

Answer 15

• In vitro means that something is being done in a test tube.
• RNA polymerase II needs transcription factors for in vitro transcription.

Question 16

What is the eukaryotic core promoter and what are the four core promoter elements? Where are they located?

Answer 16

• The core promoter in eukaryotes is where RNA polymerase II binds to.
• The four core promoter elements are:
  1. ) B recognition site located at (-35).
  2. ) TATA Box located at (-25).
  3. ) Initiator motif: transcription start site location
  4. ) Downstream core promoter located (+30)

Question 17

What are the purposes of Transcription Factors II D and H?

Answer 17

• TFIID contains the TATA Binding Protein (TBP) which is the first to bind to the TATA core promoter region. This initiates other transcription factors to come and help RNA polymerase bind to the DNA strand. The TBP distorts the DNA when it binds to the TATA box.
• TFIIH contains kinase and DNA helicase which unwinds the double-stranded DNA molecule to RNA polymerase II and provides access to the single template DNA strand. TFIIH is the last transcription factor which binds; gives the “go” signal.
• TFIIH also adds phosphate groups to the C-Terminal tail of RNAPII.

Question 18

What is the order of RNA polymerase transcription of a Prokaryotic gene?

Answer 18

1. ) Closed complex
2. ) Open complex
3. ) Initial transcribing complex
4. ) Promoter clearance