4) Control of Transcription 2

Question 1

What does the standard sigma factor do?

Answer 1

Recognises the -35 box (consensus sequence)

If a different sigma factor is used by the RNA polymerase, it will recognise a different sequence

Question 2

What do alternative sigma factors do?

Answer 2

Used in some situations for switching on and off different groups of gene sin bacteria

Example: Response to heat shock 
Genes activated by heat shock have different -35 and -10 box sequences 
Example: 
Normal promoter: TTGACA 
Normal gene: TATAAT 

Heat shock promoter: CCTTGAA
Heat shock gene: CCATNT
These can recognise these heat shock genes and are different sigma factor to the core RNA polymerase enzyme= Different recognition capabilities

ROLE: Numerous different sigma factors that help RNA polymerase move to particular genes that need transcribing under specific situations

All but one recognise specific -35 and -10 regions

Question 3

When are alternative sigma factors synthesised?

Answer 3

Not usually synthesised, but production is induced by particular conditions such as heat shock= Cause protein mis-folding

Heat shock: Alternative sigma factor (sigma 32), much smaller than normal sigma factor
They do not recognise the consensus promoter sequences but recognise the heat shock genes

These different sequences are found in promoters of heat shock genes= Genes that need to be switched on specifically in response to heat shock

Question 4

What does regulating transcription termination do? What are the 2 different types?

Answer 4

Decides whether additional genes are transcribed

1) Antiterminators
2) Attenuation

Question 5

What does antitermination do?

Answer 5

Allows terminator signals ro be ignored

Normally: When RNA polymerase encounters a terminator sequence= Stop
However: Antiterminator protein allows the terminators to be ignored

Question 6

What happens when a RNA polymerase encounters an antiterminator protein?

Answer 6

1) RNA pol moves along the promoter and reaches the antitermination site
2) RNA pol picks up the protein and brings it along with it
Having the anti terminator protein allows Pol termination signal 1 to keep transcribing the next block of genes until it reaches terminator 2= Stops

It is a way of switching on or off genes at the end of an operon

Question 7

What happens in attenuation mechanism?

Answer 7

In the trp operon:
Small deletions before the first structure gene (trpE)= Deletions in the LEADER sequence= Dramatically INCREASE the amount of transcription through the operon
INDICATES: Deletions remove a block to transcription

In wild type bacteria= No deletions= Most transcripts end before true + Rate of transcription termination at this point is regulated by the amount of free tryptophan in the growth media

Specifies the translation of a short peptide for which the amino acid is not available to construct= Can influence whether or not the terminator is made and whether or not the biosynthetic genes are transcribed

Question 8

What did DNA sequencing of the long leader region (trpL) show it has?

Answer 8

-There were 3 overlapping regions
-The 3rd (nearest trpE) overlapping region is a transcription terminator
-Since they overlap= Cannot ALL form stem and loop structures at the same time
CAN FORM:
Stem and loops for 1st and 3rd (terminator) region OR just stem and loop 2nd region= NO terminator

Question 9

How is the terminator formed?

Answer 9

• In the overlapping regions of 1 and 2= Small open reading frame which is a short gene
• An adjacent pair of the amino acids in this peptide are tryptophan
• UNLESS prevented: Stem and loops will form in order in which they are transcibed= Stem and loop 1 forms first, stopping 2nd and therefore Stem and loop 3 forms and terminates transcription

Question 10

What happens of the ORF is translated under conditions of plentiful tryptophan?

Answer 10

Same thing will happen:

• Ribosomes will follow closely behind the RNA polymerase
• End of the ORF before the 2nd stem and loop structure is transcribed
• Ribosome dissociates, the 1st stem and loop structure will form= Terminator will form

Question 11

What happens when there are reduced tryptophan concentrations?

Answer 11

• Ribosomes will stall over the trp codons
• Prevents stem and loop 1 from forming
• Result: Stem and loop 2 will be transcribed and form
• STOPS 3rd stem and loop from forming= No termination= RNA polymerase will now progress into the operon and allow its expression

Question 12

So…. What actually happens in attenuation?

Answer 12

So, in the trp operon:
Low concentrations of tryptophan= Ribosome stalls in a crucial position= Stem and loop 1 is not made= the overlapping ORF gene is not transcribed as it needs tryptophan to be synthesised

This allows stem and loop 2 to be transcribed and formed= Prevents 3rd stem and loop (terminator) from forming= No terminator= RNA polymerase can keep transcribing the operon