Prokaryotic transcription initiation

Question 1

What are the 3 stages of transcripion?

Answer 1

Initiation, elongation termination

Question 2

What stage of transcription is the rate limiting step?

Answer 2

Initiation

Question 3

In which stage of transcription does most regulation occur?

Answer 3

Initiation

Question 4

How did the initial model of transcription from the 1980s describe how transcription initiation occurred?

Answer 4

1. Sigma helps the rest of the holoenzyme bind tightly to the promoter and forms the closed complex
2. Sigma separates the two strands of DNA at the promoter and forms the open complex
3. RNAP transcribes the first two nucleotides
4. Promoter is cleared and elongation starts

Question 5

What is the closed complex?

Answer 5

dsDNA is still annealed, and sigma is making all the sequence specific binding (except for any UP elements)

Question 6

How is the closed complex converted to the open complex?

Answer 6

dsDNA is melted into ssDNA and RNAP undergoes a conformation change

Question 7

What is sigma doing during the conversion from the closed complex to the open complex?

Answer 7

Binding to the -10 sequence, then it grabs a flipped base from the non-template strand and destabilizes the double helix so the RNAP core can unwind the bases to expose the transcription start site. It will also stabilize the dsDNA/ssDNA junction

Question 8

What is the open complex?

Answer 8

dsDNA is unwound into ssDNA, and the template strand is fed into the RNAP active site, then sigma subunit gets displaced

Question 9

What is the initiating riboucleotide?

Answer 9

The first nucleotide to be incorporated into a new RNA transcript

Question 10

How is the RNA transcript initially synthesized?

Answer 10

Sigma binds to the initiating ribonucleotide, then it gets incorporated into the new strand

Question 11

What is abortive transcription?

Answer 11

A cycle done by RNAP where it synthesizes a short transcript and releases it, and keeps doing that until it achieves promoter clearance

Question 12

Is abortive transcription unique to prokaryotes?

Answer 12

No, it’s a hallmark of transcription is every organism. Prokaryote or eukaryote

Question 13

Why does abortive transcription happen?

Answer 13

Not 100% sure, but believed to be caused by sigma competing with the RNA chain in the active site

Question 14

Is RNAP moving during abortive transcription?

Answer 14

No, it’s staying stationary and sort of pulling on the DNA, synthesizing the abortive transcript, then letting the DNA and the transcript go. It doesn’t move until it achieves promoter clearance

Question 15

How does promoter clearance occur?

Answer 15

RNAP has to overcome the stable interactions with the promoter, so it can detach and move along the template DNA

Question 16

What are the 4 steps of transcription initiation?

Answer 16

1. Formation of closed complex
2. Conversion to open complex
3. Initial RNA synthesis and abortive transcription
4. Promoter clearance

Question 17

What is the sigma cycle?

Answer 17

Where sigma will associate with the RNAP core, bind to the promoter and initiate transcription, then dissociate during elongation, then go associate with another RNAP core to initiate more transcription

Question 18

Does the sigma cycle always happen?

Answer 18

No, sometimes sigma stays attached and can pause RNAP during transcription and act as another point of regulation