Chapter 15- Bacterial Cell Signaling

Question 1

Two component signaling

Answer 1

A simpler cell signaling system that is widespread in bacteria and archaea. It is a signal transduction system that enables bacteria to adapt to changes in external milieu and helps with responses to environmental stress. Similar (but more complex) systems exist in eukaryotes

Question 2

2 main components of bacterial signaling systems

Answer 2

1. Histidine kinase- a receptor in the cell membrane sensing the environmental stimulus
2. Response regulator- The signal is transmitted across the membrane and through the cytoplasm until it reaches the cytoplasmic target

Question 3

Cytoplasmic target of signaling systems

Answer 3

The cytoplasmic component will modulate transcriptional machinery. Sometimes these are enzymes, other times they can be other cellular components (like flagella). Most often, the cytoplasmic signaling protein is a transcription factor itself. When it is activated, it goes to the nucleus and activates transcription of genes that will help the bacteria to deal with the change in environment

Question 4

Histidine kinase

Answer 4

Sensor kinase that receives the signal- temp, nutrients, toxins, osmotic stress, cell density, misfolded proteins, etc. Histidine kinase may sense the signal itself or receive it from another sensor protein upstream. The kinase
autophosphorylates once it receives the signal, and this autophosphorylation activates it. It then goes on to activate the response regulator

Question 5

Response regulator

Answer 5

Partner protein to HK- it receives a phosphate group from the kinase to activate. The RR is what’s most often a transcription factor

Question 6

Histidine kinase mechanism (4 steps)

Answer 6

1. The input domain of the histidine kinase is located at the N-terminus and is what receives the signal. This part of the histidine is variable from kinase to kinase
2. The transmitter domain of histidine kinase is located at the C terminus, where there is a conserved histidine which is phosphorylated. Its function is to receive and pass the signal to the response regulator
3. When the histidine kinase senses a change in the environment, it will autophosphorylate, producing ADP
4. The histidine kinase transfers the phosphate to the response regulator

Question 7

Response regulator mechanism

Answer 7

1. Contains a receiver domain at the N-terminus that is conserved. The phosphate is transferred from histidine kinase to the aspartate on the receiver domain. The RR is then activated
2. The RR uses its output domain at the C terminus to change things in the cell. It will bind to or phosphorylate its target. It will bind to DNA if it is a transcription factor

Question 8

Phosphatase

Answer 8

A possible extra component of the two component system. Dephosphorylates and therefore deactivates the RR. “Recharges” RR so it may respond to new signal, or may silence a signal.
The phosphatase may be HK, RR itself (phosphatase domain), or a separate protein

Question 9

Phosphotransferases

Answer 9

A possible extra component of the two component system. They are several proteins that serve as a phosphorelay “bridge” from HK to RR. This is basically a bridge across which the signal can travel.

Question 10

Sensor protein

Answer 10

A possible extra component of the two component system. It is the first protein in the signal transduction pathway. Senses/responds to external stimulus. Can be HK, or it can be a separate protein upstream of HK that will sense a change in the environment

Question 11

Possible variations of the 2 component pathway (4)

Answer 11

1. The phosphate can be transferred to the response regulator and activate it
2. Phosphotransferases between HK and RR receive the phosphate. The phosphate transfers between the PT proteins and is eventually transferred to RR
3. A sensor protein is located upstream of HK. It will detect changes in environment and pass the phosphate signal to HK and then to RR (or to PT proteins if present)
4. HK is usually a membrane protein, but not always. It can be cytoplasmic, with the sensor being the only membrane protein

Question 12

Other aspects of the two component system

Answer 12

These systems do not always cause an increase in phosphorylation- the signal may result in dephosphorylation of RR. In this case, phosphatase would be another necessary component. They signal may also result in inhibition of phosphorylation of RR. In either case, there is a fall in phosphorylation of RR. Occurs when HK is bifunctional- it has kinase activity and phosphatase activity

Question 13

Aerobic respiratory control (Arc) system

Answer 13

Responds to a shift of bacteria to an anaerobic environment- when it moves from an environment with oxygen to an environment without oxygen. Its HK is called ArcB and its RR is called ArcA

Question 14

What changes occur in the aerobic respiratory control (Arc) system (4)

Answer 14

1. Repression of genes for citric acid cycle enzymes upon shift to an anaerobic environment. A full citric acid cycle is not needed if there is no oxygen
2. Induction of gene for pyruvate formate-lyase
3. Induction of gene for cytochrome d oxidase.
4. Results in non-cyclic citric acid metabolism

Question 15

Pyruvate formate-lyase

Answer 15

Converts pyruvate to acetyl-CoA in lieu of pyruvate dehydrogenase

Question 16

Cytochrome D oxidase

Answer 16

Cytochrome oxidase is where the final electron acceptor (oxygen) is located for the ETC. However, in the absence of oxygen, a different enzyme called cytochrome D oxidase is synthesized. Cytochrome D oxidase has a much higher binding affinity for oxygen than cytochrome C oxidase does. This ensures that any of the small amount of oxygen present in an anaerobic environment can be used

Question 17

Non-cyclic citric acid cycle

Answer 17

Occurs in bacteria in anaerobic environments. This pathway splits into 2 linear, parallel pathways. The product of one side is succinyl CoA and the product of the other side is alpha ketoglutarate. The citric acid cycle is still partially functional in an anaerobic environment so the intermediates of the pathway can serve as building blocks for organic molecules

Question 18

Sensors of the Arc system (4)

Answer 18

1. Reduced e- transport carrier (such as a quinone)
2. Pyruvate
3. Acetate
4. D-lactate

Question 19

Why are the 4 molecules used as sensors for the Arc?

Answer 19

Reduced electron transport carriers like quinone indicate that there is less ETC activity as a result of low oxygen. Pyruvate,
acetate, and D-lactate will be increased when oxygen is lacking. Pyruvate will be increased because there is less of a citric acid cycle. Lactate increases because when oxygen is limited, bacteria will partially switch to fermentation to produce energy

Question 20

Lactate fermentation (2)

Answer 20

1. NADH and pyruvate feed into this pathway. There is a donation of 2 hydrogens from NADH, which creates lactate
2. These 2 hydrogens are added to 2 pyruvate molecules, which gives us 2 molecules of lactate

Question 21

2 component Arc system mechanism (3 steps)

Answer 21

1. ArcB is a membrane protein that acts as the HK. It detects signals like reduced electron transport carriers, increased pyruvate, and others
2. The presence of these signals activates (autophosphorylates) ArcB
3. The phosphate is then transferred to ArcA. Arc acts as a transcription factor that will transcribe pyruvate formate lyase, cytochrome D oxidase, and reductive enzymes for the citric acid cycle (what modifies the citric acid cycle to make it noncyclical)

Question 22

N-terminus

Answer 22

The first part of the protein that exits the ribosome during protein biosynthesis

Question 23

C-terminus

Answer 23

C-terminus is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). It is the opposite end from the N-terminus

Question 24

Anaerobic respiration

Answer 24

When organisms have something different from oxygen as their final electron acceptor

Question 25

Nar system

Answer 25

Allows utilization of nitrate or nitrite (nitrogen, really) as an e- acceptor during anaerobic respiration. It is a dual 2 component system as there are 2 histidine kinases (NarX and NarQ) and 2 response regulators (NarL and NarP). Therefore this system is essentially 2 systems. The NarX system allows for the utilization of nitrate and the NarQ system allows for utilization of nitrite

Question 26

NarX system in the presence of nitrate (3 steps)

Answer 26

In the presence of nitrate:
1. NarX autophosphorylates-it is a histidine kinase
2. NarX transfers the phosphate and phosphorylates NarL (the response regulator)
3. P-NarL stimulates transcription of narG operon- these are nitrate reductase genes

Question 27

Nitrate reductase genes

Answer 27

Genes that encode proteins that allow the bacteria to take nitrate and utilize it as the final electron acceptor

Question 28

NarX system in the presence of nitrite (3 steps)

Answer 28

1. NarX (HK) acts as phosphatase
2. NarX dephosphorylates NarL (response regulator)
3. No stimulation of narG operon, so the nitrate reductase genes are not expressed

Question 29

NarQ system in the presence of nitrite (3 steps)

Answer 29

1. NarQ (HK) autophosphorylates
2. NarQ transfers its phosphate and phosphorylates NarP (RR)
3. Phosphorylated NarP stimulates transcription of the nrfA operon- contains periplasmic nitrite reductase genes, allowing the bacteria to use nitrite as the final electron acceptor

Question 30

NarQ system in the presence of nitrate

Answer 30

NarQ acts as a phosphatase

Question 31

Assimilatory metabolism of nitrogen definition

Answer 31

Using nitrogen from the environment to build molecules, and energy is released as molecules are built in the cell. These nitrogen are being assimilated into the molecules being built in the bacterial cell.

Question 32

Assimilatory metabolism of nitrogen 2 component system

Answer 32

This 2 component system acts as a response to the supply of nitrogen. There are multiple sources of nitrogen. It can be inorganic (nitrate, N2) or organic (amino acids, urea). Regardless of the form, all nitrogenous compounds are reduced to ammonia by bacteria. Ammonia is then utilized as the nitrogen source

Question 33

2 reactions of enteric bacteria for the assimilation of ammonia

Answer 33

The 2 component system of nitrogen assimilation will work in enteric (GI tract) bacteria. They have 2 reactions that allow for assimilation of ammonia into the cell
1. Reactions that produce the amino acid glutamate, which needs α-ketoglutarate as a precursor
2. Reactions that produce the amino acid glutamine, which needs glutamate as a precursor

Question 34

Assimilatory metabolism of nitrogen reactions- reactants and products

Answer 34

1. When there is a lot of ammonia and α-ketoglutarate is present, the cell can make glutamate
2. Glutamate can also react with ammonia to form glutamine

Question 35

Ntr regulon

Answer 35

Ammonia is the preferred source of nitrogen, so this is what the 2 component system will detect. It represses genes that are needed for the assimilation of alternative nitrogen compounds. When limited, there is induction of genes for production of ammonia from alternative sources (aka other sources of nitrogen). The Ntr regulon consists of several non-contiguous operons that regulates assimilation of alternative sources of nitrogen

Question 36

Ntr regulon system- high ammonia levels

Answer 36

If we have high ammonia levels, the cell will sense that due to rising glutamine levels. This makes sense because if there is a lot of ammonia, the nitrogen assimilation reactions can just keep going and form glutamate and glutamine. In this case, the transcription of the Ntr regulon of genes that allows for utilization of an alternative source of nitrogen is repressed.

Question 37

Ntr regulon system- low ammonia levels

Answer 37

Low ammonia levels results in an increase in α-ketoglutarate. This is because if ammonia is limited, the nitrogen assimilation reactions cannot be carried out. α-ketoglutarate increases because it cannot be converted to glutamate. In this case, Ntr transcription is activated. This allows for the assimilation of other sources of nitrogen

Question 38

Ntr system mechanism when ammonia is low (5 steps)

Answer 38

1. NR2 is the histidine kinase and P2 is bound to it in the resting state. When ammonia is low, α-ketoglutarate concentration increases in the cytoplasm
2. Increased α-ketoglutarate causes the P2 protein to unbind from NR2
3. At this point, the HK is activated so NR2 is phosphorylated
4. NR2 transfers its phosphate to the response regulator (NR1)
5. This causes the transcription of the Ntr regulon, which allows the bacteria to use an alternative source of nitrogen

Question 39

Ntr system mechanism when ammonia is high (3 steps)

Answer 39

1. When ammonia is high, glutamine increases in the cytoplasm. This causes the P2 inhibitory protein to bind to NR2 (the HK)
2. The HK’s phosphatase activity is stimulated by P2 binding activity. That ensures that there are not phosphates on the response regulator
3. The Ntr regulon is repressed because there is no need to deal with alternative nitrogen sources

Question 40

Pho regulon

Answer 40

Found in E. coli and is a response to phosphate supply. When the amounts of phosphate are low, E. coli stimulates transcription of 38 genes involved in phosphate assimilation. The 2 component system regulating the pho regulon is called the IM phosphate uptake system (Pst). This system has many extra components. The regulon contains 14 genes for uptake & breakdown of phosphonate

Question 41

Components of the IM phosphate uptake system (Pst)

Answer 41

1. Periplasmic alkaline phosphatase (PhoA)- generates phosphate from organic phosphate esters
2. OM porin channel (PhoE)-allows the uptake of anions, including phosphate

Question 42

Regulation of the Pho regulon when phosphate is high

Answer 42

When Pi is high in the environment, the Pho regulon is repressed. In this situation, the high amount of phosphate is transported into the cell by a low affinity transporter called Pit

Question 43

Regulation of the Pho regulon when phosphate is low

Answer 43

Transcription of the Pho regulon is activated, which increases the expression of proteins allowing for the uptake of phosphate. The uptake of phosphate is necessary since it is low in the environment

Question 44

PhoR

Answer 44

The histidine kinase in the pho system

Question 45

Pho system mechanism- high levels of phosphate (3 steps)

Answer 45

1. In a system where there is plenty of inorganic phosphate, phosphate will bind to the PstS protein, which will in turn bind to a complex of proteins called PstA, PstB, and PstC.
2. In that situation, the PhoR HK becomes bound to the PstABC complex. These proteins form a locked complex
3. PhoR acts as a phosphatase, and its response regulator (PhoB) is connected to it. This inhibits the signal from getting to PhoB

Question 46

Pho system mechanism- low levels of phosphate (4 steps)

Answer 46

1. In this situation, there is not enough phosphate to bind PstS. This means that the HK will not be locked in a complex.
2. The HK will be free to dimerize with another HK. Once the 2 HKs dimerize, they autophosphorylate. There are 2 phosphates, one on each subunit
3. The phosphates are bound transferred to the attached PhoB subunits (the RR)
4. The PhoB subunits are activated and go to transcribe the pho regulon. This allows for increased uptake of phosphate into the cell as phosphate is limited

Question 47

Rhodobacter capsulatus

Answer 47

Purple nonsulfur photosynthetic bacteria. They grow with O2 and derive energy from aerobic respiration. However, if they are shifted to an oxygen limiting environment, they grow anaerobically and derive energy from photosynthesis. Their photosynthetic genes are regulated at the level of transcription in multiple different ways

Question 48

3 ways of regulating bacterial photosynthetic genes at the level of transcription

Answer 48

1. Repressor activated by O2
2. Inducer activated by lack of O2
3. Inducer activated by low light

Question 49

Purple photosynthetic bacteria at high levels of oxygen (4 steps)

Answer 49

1. Oxygen enters the bacterial cell and oxidizes a protein called CrtJ. This causes CrtJ to form a disulfide bond, changing the structure of the protein
2. The protein will not be able to bind to its promoter region
3. Repression of photosynthetic genes occurs, as the protein is unable to activate transcription. There is plenty of oxygen in the environment, so there is no need to obtain energy through photosynthesis
4. This part of the system ensures that photosynthesis only occurs under oxygen limiting conditions

Question 50

In photosynthetic bacteria, what do the photosynthetic genes code for?

Answer 50

Codes for biosynthetic proteins for pigment synthesis, as well as proteins of the reaction center and light harvesting complexes

Question 51

Anaerobic induction

Answer 51

A two-component system that stimulates transcription of genes for light-harvesting & reaction center complexes. This system only has 2 components- RegB (HK) and RegA (RR). RegB autophosphorylates under anaerobic conditions. RegA is phosphorylated by RegB, and it activates transcription of the photosynthetic genes

Question 52

Anaerobic induction mechanism (normal light levels) (3 steps)

Answer 52

1. Anaerobic conditions activates RegB (the HK). RegB autophosphorylates
2. RegB transfers its phosphate to RegA (the RR)
3. RegA induces transcription of the genes that are needed for photosynthesis

Question 53

Anaerobic induction mechanism at low light (3)

Answer 53

1. Response to low light in an anaerobic environment- photosynthesis must be increased in this case
2. A protein called HvrA is a transcription factor that helps to transcribe certain photosynthetic pigments. Transcription of this transcription factor increases in low light, so there are more photosynthetic pigments
3. Therefore, even at low light, the bacteria can still take in light energy

Question 54

E. coli response to osmotic pressure and temperature in freshwater environments

Answer 54

In freshwater, there are low levels of solutes and low temperatures. In this case, a large OmpF channel is made. This is meant to increase the uptake of solutes- it allows the solutes into the cell since initially there are less solutes in the environment

Question 55

E. coli response to osmotic pressure and temperature in intestinal environments

Answer 55

The intestines have higher levels of solutes, like sodium chloride, and higher temperatures, ranging from 30-37 degrees. In this case, a small OmpC channel is made. A smaller pore slows down the uptake of molecules. It will also slow down the flow of toxic substances, since there are many of those in the gastrointestinal track

Question 56

Regulation of OmpF and OmpC

Answer 56

A two component system is involved here. The HK is EnvZ- it autophosphorylates in high osmolarity conditions, and low osmolarity conditions prevent kinase activity. The RR is OmpR, and it is phosphorylated by EnvZ. The RR acts as an activator of transcription of OmpC (the smaller pore) and as a repressor of transcription of OmpF (the larger pore).

Question 57

E. coli response to osmotic pressure mechanism (high osmolarity)

Answer 57

1. In a high osmolarity environment, EnvZ will autophosphorylate
2. It will transfer the phosphate to OmpR (the RR)
3. This causes transcription of OmpC and repression of OmpF. It is a smaller pore that will slow down the flow of toxic substances into the cell

Question 58

E. coli response to osmotic pressure mechanism (low osmolarity)

Answer 58

1. In a low osmolarity environment, EnvZ phosphatase activity is activated.
2. This ensures that there is no phosphorylation of the RR
3. Transcription of OmpF is activated, OmpC transcription is repressed. OmpF is the larger pore and is necessary for solute uptake in this type of environment