2. control of transcription termination of prokaryotic cells

part II of prokaryotic gene expression regulation

2. How is transcription termination regulated = attenuation control 

By different positions of terminators

if first terminator at the beginning/first —> small mRNA as RNA pol stops at term1

if terminator father down at the end the coin sequence —> mRNA coding for gene is transcribed

2. what is the stricture of an independent Rho terminator

interrupted palindrome (sequence above is the same as complementary sequence), followed by a T/A rich sequence

• RNA pol transcribes palindrome sequence

• complementary sequence hybridise to form a stem loop sequence upstream of T/A rich sequence

• RNA pol pause due to this structure at the T/A rich sequence

• RNA pol has a weak interaction w/ T/A rich sequence (no C/G) —> immutable interaction

• termination os transcription 

2. how can termination be prevented ?

by preventing the formation of the stem loop structure upstream of the T/A rich sequence, which can be blocked by : 

• translation of a leader protein (ribosome mediated attenuation )

• w/ a protein

• w/ an tRNA = box riboswitch  

• by binding a metabolite = riboswitch

2.1 control of termination by a leader protein, give an e.g 

pyr operon in e.coli

the operon codes for pyrimidine bases (UTP) in RNA 

the operon had 2 terminators Term1 and Term2

• at the beginning of the operon there is a sequence that’s codes for 

2.1 how is the control of termination of the pyr operon regulated in the presence of many UTPs

term 1 is used when there’s lots of UTPs (as less pyr bases are needed)

• translation of leading protein before RNA is complete w/ transcription

• RNA pol adds quickly the complementary U bases to the T/A rich sequences 

• RNA forms a terminator stem loop structure upstream of U rich region 

• termination occurs at term1 —> RNA pol falls off and theres no expression of genes encoding for pyr bases

2.1 how is the control of termination of the pyr operon regulated in the presence of little UTPs

term 2 is utilised as more pyr bases are needed : UTP affects transcription efficiency

• the ribosomes translating the leading proteins remains stuck behind the RNA pol

• no stem loops structure

• RNA pol is slowly adding U complementary bases to the T/A rich sequences 

• RNA pol is not paused and continues transcription —> gene encoding for pyr bases are expressed 

2.1 give me a quick summary of how pyr operon termination is controlled by a leader protein 

• High UTP → efficient transcription/translation → Term1 stem-loop forms → transcription terminated → no pyr gene expression.

• Low UTP → stalled ribosome + slow polymerase → no Term1 → transcription continues → pyr genes ON.

2.2 control of termination w/ a protein, give me an e.g

the hut operon in B.subtilis

hut = histdine utilisation ( codes for enzymes that break down His and produce ATP)

2.2 what protein regulates the termination of hut operon

HutP = RNA binding protein = regulatory protein

2.2 what happens to the termination of the hut operon in the presence of little His

HutP is inactive —> doesn’t bind to RNA

termi ator stem loop structure is formed upstream of the U rich region

detachment of RNA pol 

termination of transcription —> no gene expressed

2.2 what happens to the termination of the hut operon in the presence of many His

His binds to HutP

• HutP can bind to specific sequence CAG/UAG on RNA 

HutP binds upstream/overlapping palindromic sequences 

Inhibition of the formation of terminator stem loop structure 

RNA pol doesn’t terminate at term 1 —> expression of genes 

2.2. what are the alternative RNA structures to control termination

terminator stem loop structure —> two palindromic sequences hybridise upstream of the U rich region

anti-terminator stem loop structure —> two palindromic sequences hybridise before a sequence upstream of U rich region

2.2. what’s another example of termination control w/ a protein (also controlled by leader protein)

pyr operon that encodes for different pyr genes :

• pyrR : codes for protein that regulates pyr operon expression according to the level of UTPs

• pyrP : encodes for uracil permeate

• pyrB & following genes : codes for enzymes involved in UMP (pyrimidine nucleotide) synthase

2.2 how is termination of pyr operon controlled in low levels of UTP/UMP 

PyrR is inactive and doesn’t bind to RNA

anti terminator structure (S2-S3) is very stable (secondary structure of RNA )

RNA pol doesn’t terminate and whole pyr operon is expressed 

2.2 how is termination of pyr operon controlled in high levels of UTP/UMP

UMP/UTP binds to PyrR —> dimerisation —> active 

PyrR binds to RNA and induces stem loops structure of S1-S2 = anti-anti terminator stem loops 

S3-S4 hybridise and forms a terminator stem loops 

Termination at the attenuators 

2.2 what happens the termination control if there’s lots of UMP/UTP but even more GMP/GTP

competition between GT/MP and UT/MP to bind to PyrR

• when UT/MP binds to PyrR —> active form

• when GT/MP binds to PyrR —> inactive form 

  • PyrR doesn’t bind to RNA, formation of anti terminator stem loop structure 

  • no termination and pyr operon expressed

TO KEEP EQUILIBRIUM OF PYRIMIDINE/PURINE

2.3 what is the definition of a riboswitch

cis regulatory system (regulation from same sequence/DNA) of an mRNA that directly senses physiological signals causing a change in the RNA structure

• terminator stem loop

• anti terminator stem loop

riboswitch can be a : 

• metabolite e.g sugar, vitamin

• t box : tRNA 

2.3 what is an example of a box riboswitch

gene encoding for tyr synthetase tRNA

2.3 how does the ration of loaded to unloaded tRNA influence the regulation of certain genes

if there is an excess of uncharged tRNA :

• increased expression of genes involved in the biosynthesis of the corresponding AA

• increased expression of the gene encoding for the corresponding aminoacyl transferase

2.3 using the example of tyrosine how is the tRNA loaded with tyr

All tRNA have a CCA sequence at their 3’ = AA attachment site

tyrosyl-tRNA synthetase (aminoacyl tRNA synthetase) will charge the tRNA w/ tyr at the CCA sequence 

loaded tRNA can bind to codon and be used for translation 

2.3 what does the tyre operon encoding for tyrosyl-tRNA synthetase look like

contains 2 terminators :

• term1 after the promoter —> used when there’s high ratio of charges tRNA so less tyrosyl-tRNA synthetase needed 

• term2 at the end of the tyrS gene —> used when there’s a high ratio of uncharged tRNA so more tyrosyl-tRNA synthetase is needed 

2.3 if there’s few uncharged tRNA/many charged tRNA what is the termination control for tyrS

mRNA transcribed has a:

• stem loop structure w/ codon recognised by corresponding tRNA ( UAC for tyr codon) —> remains unbound to tRNA

• T box which contains an anti terminator + UGG triplet doesn’t bind to S1 —. no anti terminator loop structure formed

• Terminator stem loop of S1-S2 formed at the U rich region 

—> termination at term1 as there’s no uncharged tRNA to bind to t-box

2.3 if there’s many uncharged tRNA/few charged tRNA what is the termination control for tyrS

mRNA transcribed :

• the uncharged tRNA binds to codon (UAC) on the stem loop structure and the UGG triplet on the T-box

  • stabilises the formation of an anti terminator team loop structure between T box and S1

• no terminator stem loops structure 

• elongation continues 

—> termination at tem 2 and expressie of tyrS