week 3 - global regulation

Question 1

local vs global regulation

Answer 1

• considered single operons
  o lac operon
  o trp operon
• encode only a few genes
  o all that is needed to a very specific purpose
• respond to a simple signal (usually a small molecule)
• examples of local regulation

Question 2

sometimes multiple genes are needed to change expression

Answer 2

o this enables e.g. e. coli to survive in many different environments
- shift from aerobic to anaerobic growth
- change in external pH
- osmotic shock DNA damage
- change in temperature
- change in level of available glucose
o glucose levels drop, cAMP levels rise (this activates CRP protein)
o CRP protein binds cAMP
o Can bind and activate many different operons (not just lac)
o CRP Is acting as a global regulator
 Regulates many different operons

Question 3

operon

Answer 3

• Set of genes, transcribed as a single mRNA
• Adjacent on DNA

Question 4

regulon

Answer 4

• Set of genes and operons regulated by the same protein
• May be scattered across whole genome

Question 5

Regulon example

Answer 5

LexA

• Genes of the LexA regulon are for repair of DNA damage
• All are normally repressed by LexA protein

Question 6

Regulon example
LexA
what does it do

Answer 6

• Damaged DNA forms a complex with RecA proteins and activates it
  o RecA: Binds single stranded RNA, Changed shape, Now RecA binds to LexA
• Activated RecA cleaves LexA
• Genes which are normally repressed by LexA are now expressed
• Genes of the LexA regulon are for repair of DNA damage
• All are normally repressed by LexA protein

Question 7

Regulon example
LexA
What will happen to the LexA regulon if recA is deleted?

Answer 7

• Mutation recA strain cannot respond to UV radiation

The cell can’t mount an SOS response.

It becomes hypersensitive to DNA damage (e.g., UV light, certain antibiotics).

DNA damage will accumulate, potentially leading to cell death

Question 8

GLOBAL REGULATION
How widespread?

Answer 8

• About 10% of genes in e. coli are regulators
• 50% of all genes in e coli are controlled directly or indirectly by one or more of seven global regulators

Question 9

GLOBAL REGULATION
Why does it occur?

Answer 9

• Requires fewer regulators
  o More efficient use of resources
• Enables co-ordinated response to major changes in growth conditions
• Enables different combinations of key operons to be expressed under different conditions

Question 10

GLOBAL REGULATION
How does it happen?

Answer 10

• Many mechanisms, but all have in common
  o Something has to detect the signal or change in conditions
  o This has to be linked somehow to altered gene expression

Question 11

GLOBAL REGULATION
examples

Answer 11

aerobic/anaerobic switch

heat shock

Question 12

GLOBAL REGULATION
Aerobic/anaerobic switch

Answer 12

• E coli can grow aerobically or anaerobically (facultative anaerobe)
• If switch to anaerobic growth need to:
  o Repress expression of genes involved in aerobic pathways (e.g. TCA cycle, FA oxidation)
  o Repress expression of genes involved in protection against oxygen and its biproducts
  o Repress expression of expensive genes where proteins require large amounts of ATP

Question 13

GLOBAL REGULATION
Aerobic/anaerobic switch
what is responsible for the switch

Answer 13

o ArcAB system is an example of a two-component system

sensor kinase senses changes in conditions
(autoposphorylates itself and passes phosphate onto a response regulator)

ArcB stimulated to become active by lactate, pryuvate, acetate. ArcA is then phosphorylated

represses a wide range of genes (TAC cycle; cirtrate synthase, operons involved in fatty acid oxidation)

also activates some genes

this represses aerobic respiration

Question 14

GLOBAL REGULATION
Aerobic/anaerobic switch
What phenotype would you predict for an arc ArcA or arcB deletion?

Answer 14

• No repression of aerobic respiration

Question 15

GLOBAL REGULATION
Aerobic/anaerobic switch
ArcB

Answer 15

Sensor kinase (membrane-bound) that detects low oxygen (reducing conditions)

Question 16

GLOBAL REGULATION
Aerobic/anaerobic switch
ArcA

Answer 16

Response regulator (cytoplasmic); when phosphorylated, it represses or activates gene expression

Question 17

GLOBAL REGULATION
Aerobic/anaerobic switch
Aerobic conditions

Answer 17

ArcB is inactive → no phosphorylation of ArcA.

ArcA remains unphosphorylated.

Genes for aerobic respiration stay on (e.g., TCA cycle, cytochrome oxidases).

Anaerobic genes are not repressed by ArcA.

Question 18

GLOBAL REGULATION
Aerobic/anaerobic switch
anaerobic conditions

Answer 18

ArcB senses reducing conditions and autophosphorylates.

ArcB transfers the phosphate to ArcA → ArcA~P (phosphorylated form).

ArcA~P acts as a transcriptional regulator:

Represses genes for aerobic respiration (e.g., sdh, cyo).

Activates genes for anaerobic pathways (e.g., cyd, fermentation genes).

Question 19

GLOBAL REGULATION
heat shock

Answer 19

• For e coli: going from 37 degrees to 43
  o To do with folded protein structures (on the edge of stability) most organisms have evolved to grow at a restricted temperature range
• Nearly all organisms show heat shock
  o Temperature depends on their normal growth temperature
• Major consequence of heat shock is that proteins begin to unfold and to aggregate

Question 20

GLOBAL REGULATION
heat shock
genes of heat shock response

Answer 20

 Assist proteins to refold (molecular chaperones)
 Old unfolded proteins in stable, non-aggregating state until heat shock ends (molecular chaperones)
 Degrade proteins that cannot be refolded (proteases)

Question 21

GLOBAL REGULATION
heat shock
how is heat shock response regulated in e. coli

Answer 21

Do cells detect increase in temperature or presence of unfolded proteins
o Key feature: alteration of a component of RNA polymerase

Question 22

GLOBAL REGULATION
heat shock
RNA polymerase composition

Answer 22

• Alpha2betabeta
  o core enzyme contains enzymatic machinery for mRNA synthesis
   change the RNA polymerase
  o to recognise promoters and transcribe DNA, needs another factor
   sigma factor
  o e coli has several different sigma factors

Question 23

GLOBAL REGULATION
heat shock
how are heat shock genes turned on

Answer 23

heat shock genes can be turned ON by increasing levels of sigma 32

Question 24

GLOBAL REGULATION
heat shock
how is sigma 32 regulated
- post-transcriptionally

Answer 24

Heat shock causes more efficient translation of rpoH mRNA to make more sigma 32

• Sigma 32 can be regulated by its own RNA
• RNA’s Secondary structure keeps protein from being translated
• If increase temp break down the secondary structure
• Therefore more expression of Sigma 32

Question 25

GLOBAL REGULATION heat shock how is sigma 32 regulated - post-transcriptionally regulation is..

Answer 25

cis - The regulatory element in the mRNA regulates the downstream gene(s) and ONLY the downstream gene(s) - How might you test whether a different gene is regulated by a similar mechanism

Question 26

GLOBAL REGULATION heat shock how is sigma 32 regulated - post-translationally

Answer 26

o Stability of sigma 32 is regulated by molecular chaperone DnaK and protease FtsH  It is unstable in conditions where it is not needed

Question 27

GLOBAL REGULATION heat shock how is sigma 32 regulated - post-translationally UNDER NORMAL GROWTH CONDITIONS

Answer 27

very low levels of sigma32 DnaK: a molecular chaperone that helps proteins fold bound to sigma 32 DnaK delivers sigma 32 to FtsH (a protease) that degrades sigma 32

Question 28

GLOBAL REGULATION heat shock how is sigma 32 regulated - post-translationally EXPOSED TO HEAT SHOCK

Answer 28

heat shock causes proteins to unfold and DnaK bind to them (to help them refold) this competes with sigma 32 more free sigma 32 high levels of sigma 32, high transcription of heat shock genes - Sigma 32 stable and can bind to RNA polymerase - Get high levels of transcription

Question 29

GLOBAL REGULATION heat shock how is heat shock regulated

Answer 29

- Cells detect both the increase in temp and presence of unfolded proteins

Question 30

GLOBAL REGULATION heat shock how is heat shock regulated - raised temp.

Answer 30

o Increases translatability of rpoH mRNA  More sigma 32 made o Post transcriptional regulation

Question 31

GLOBAL REGULATION heat shock how is heat shock regulated increased unfolded protein

Answer 31

o Reduced DnaK binding to sigma 32 o Reduced degradation of sigma 32 by FtsH o Post translational regulation

Question 32

GLOBAL REGULATION heat shock predicted phenotypes of deletion mutations - rpoH

Answer 32

no increase in sigma 32, no post-transcriptional regulation Without σ32, the heat shock regulon cannot be activated. Consequently, heat shock proteins (like DnaK, GroEL, etc.) will not be expressed in response to temperature stress, making the strain highly sensitive to heat and other stresses that would normally activate the heat shock response. the mutant would show increased sensitivity to heat shock and poor recovery from stress because heat shock proteins cannot be synthesized.

Question 33

GLOBAL REGULATION heat shock predicted phenotypes of deletion mutations FtsH

Answer 33

Increased sigma 32 (no degredation) Normally, FtsH helps regulate σ32 levels by degrading excess σ32, ensuring that it doesn't accumulate unnecessarily. In the absence of FtsH, σ32 levels would remain abnormally high, leading to excessive transcription of heat shock genes even under normal conditions. The mutant would show elevated basal levels of heat shock proteins (due to increased σ32), which could lead to growth defects or stress response imbalances in non-stressful conditions. Under heat shock, it might show some overreaction to stress, leading to an inefficient stress response.

Question 34

GLOBAL REGULATION heat shock predicted phenotypes of deletion mutations DnaK

Answer 34

Increased sigma 32 (as reduced binding to sigma 32 so less competition) Without DnaK, less σ32 would be bound, reducing competition for binding to RNA polymerase. This would result in an increase in free σ32 available for transcription activation, leading to increased transcription of heat shock genes. The mutant would show increased levels of heat shock proteins (due to increased free σ32), similar to the FtsH deletion, but the mechanism is different. This could also lead to growth defects or an inefficient stress response.

Question 35

what is global regulation chatgpt

Answer 35

Global regulation refers to the coordinated control of multiple genes or operons across the genome by a single regulator or regulatory system. These genes may be involved in unrelated processes but are activated or repressed together in response to a shared environmental or internal signal. 🔹 Example: The LexA regulon, which controls DNA damage repair genes scattered across the chromosome. 🔹 Example: CRP, which regulates many catabolic operons in response to glucose availability.

Question 36

why does global regulation occur chatgpt

Answer 36

It allows bacteria to quickly adapt to major environmental changes (e.g., oxygen levels, temperature, nutrient availability). It's efficient: instead of regulating each gene individually, a single regulator can control many genes. Ensures a coordinated physiological response, so related pathways are turned on or off together. Saves resources by avoiding unnecessary gene expression. 🔹 Example: In oxygen-poor conditions, E. coli represses energy-expensive aerobic pathways and activates anaerobic ones via ArcAB.

Question 37

how does global regulation occur chatgpt

Answer 37

Global regulation typically involves: Sensing a signal or environmental change. Transducing that signal to a regulator protein. Regulator acts (often a transcription factor or sigma factor) to alter transcription of multiple genes.

Question 38

common mechanisms of global regulation Two-component systems chatgpt

Answer 38

A sensor kinase detects a signal and phosphorylates a response regulator ArcB/ArcA (O₂ switch)

Question 39

common mechanisms of global regulation Sigma factor switching chatgpt

Answer 39

Changing RNA polymerase’s sigma factor to recognize different promoters σ32 (heat shock)

Question 40

common mechanisms of global regulation Allosteric activation chatgpt

Answer 40

A small molecule activates a transcription factor CRP-cAMP (glucose starvation)

Question 41

common mechanisms of global regulation Protein cleavage chatgpt

Answer 41

Signal leads to cleavage of a repressor LexA-RecA (DNA damage)

Question 42

what makes a good global regulator chatgpt

Answer 42

A global regulator is typically: Responsive to a key environmental cue Capable of binding multiple promoters Often able to coordinate opposing pathways (turning one on, another off)