Lecture 4 - Exam 5

Question 1

Global regulators have the ability to ?

Answer 1

Regulate genes and operons that belong to different metabolic pathways.

Question 2

What is a regulon?

Answer 2

Often a group of genes that are involved in response to an environmental stimulus. Multiple genes and/or operons regulated by the same transcriptional regulator.

Question 3

There are multiple levels of gene regulation:

Answer 3

Transcription (most important in bacteria), post-transcription, translation, post-translation

Question 4

What are the steps in environmental sensing and response?

Answer 4

1) Recognition of environmental stimuli
2) Transduction of information
3) Induction of response via gene expression or other mechanism.

Question 5

What are the types of environmental sensing and response systems?

Answer 5

Two-component regulatory systems:
-major mode of environmental sensing and signal transduction mechanism in Bacteria and Archaea
-also found in some protozoa and plants but NOT found in mammals
Quorum sensing:
-bacterial communication system for coordinated gene regulation depending on cell density
-can be intraspecies and interspecies
Other mechanisms:
-alternative sigma factors/transcriptional regulators
-riboswitches

Question 6

Two-component regulatory systems are ideally suited to function in the _____________.

Answer 6

Integration of multiple signals.

Question 7

Different two component systems can process multiple signals into a particular response by ?

Answer 7

Interacting with one another (cross-talk or reciprocal regulation)
-and/or by phosphorylating from small molecular weight phosphodonors

Question 8

Do many two-component regulatory systems exist in bacteria?
Multiple regulatory systems are constantly functioning ___________ in generalist bacteria.

Answer 8

Yes
simultaneously

Question 9

The average number of two component regulatory systems in a bacterial genome has been estimated at around _____, with the largest number being over _____.

Answer 9

30;200

Question 10

What kind of bacteria have a greater number of two component regulatory systems and why?

Answer 10

Free-living bacteria because they have to adapt to many different environmental conditions.

Question 11

Why kind of bacteria have no two-component systems?

Answer 11

Typically these bacteria are endosymbionts or pathogens.

Question 12

General model of two component regulatory systems:
Sensor = ?
Transducer = ?

Answer 12

Signal sensed by sensor, phosphorylation of sensor and transducer, phosphorylated transducer activates transcription.
Sensor = Histidine kinase domain to sense the extracytoplasmic environment*
Transducer = Response regulator ; usually a DNA binding protein**

Question 13

All two component regulatory systems involve the sensor transmitting the signal to the response regulator, this is typically by?

Answer 13

Phosphorylation

Question 13

When an extracellular signal is received by a histidine kinase, the signal can result in three distinct outcomes:

Answer 13

1. Autokinase activity resulting in autophosphorylation of the HK.
2. Phosphotransfer to the response regulator (often acts as a transcription factor which activates or represses a particular set of response genes)
3. Phosphatase activity, leading to the dephosphorylation of the response regulator.

Question 14

Two component regulator systems consist of a sensor protein (e.g. histidine kinase) that ?

Answer 14

Transfers a high-energy phosphoryl group to the response regulator, which is often a transcription factor.

Question 15

What happens when the membrane bound sensor kinase sense a particular environmental stimulus?
-What does the kinase activity depend on?
-What happens to the phosphate group?

Answer 15

The conserved histidine residue (H) in the cytoplasmic sensor domain of this protein is phosphorylated.
-Depends on ATP which is bound to the catalytic domain
-The phosphate group is then transferred to the conserved aspartate residue (D) in the receiver domain of the cognate response regulator.

Question 16

What are the two types of two component regulatory systems?

Answer 16

Sensor-Response Regulator
Phosphorelay System

Question 17

Give an overview of sensor-response regulator two component system.

Answer 17

-The prototypical two-component system is comprised of a single sensor kinase (SK) and a single response regulator (RR)
-The input domain of the SK recognizes a specific signal from the environment.

Question 18

Describe the mechanism of the sensor-response regulator system.

Answer 18

-The sensor kinase autophosphorylates a conserved histidine residue in the kinase domain, in response to environmental signal sensing in the sensing domain.
-The autokinase reaction uses ATP to phosphorylate a conserve His residue of the Kinase domain.
-The phosphotransfer reaction transfers the phosphoryl group from the His residue of the sensor histidine kinase (SHK) to a conserved Asp residue of the REC domain of the response regulator.
-The transfer of the high energy phosphoryl group to the conserved aspartate residue in the receiver domain of the response regulator results in enhanced DNA-binding activity.

Question 19

Give an overview of the phosphorelay system.

Answer 19

-Phosphorelay systems are comprised of an SK and a terminal RR.
-These systems also contain an intermediate regulator (connector) lacking an output domain and a phosphotransfer protein with a conserved histidine for phosphorylation.
(other SHKS w/ intramembrane sensors and cytoplasmic sensors may also be present.

Question 20

Describe the phosphorelay system.

Answer 20

Are more complex and involve multiple phosphorylation steps prior to producing a response.
The His-Asp-His-Asp three step phosphorelay consists of:
A. The transfer of the phosphoryl group from the conserved histidine in the sensor kinase.
B. To aspartate and histidine residues in auxiliary proteins (connector and phosphodonor)
C. Before transfer to the aspartate in the receiver domain of the response regulator.
-Added complexity provides for multiple regulatory checkpoints and points of integration between signaling pathways.

Question 21

Most HKs are _______________.
and function as ____________.

Answer 21

Periplasmic membrane receptors.
Homodimers: autophosphorylation is a bimolecular event.

Question 22

What are the domains od the HKs?

Answer 22

-Sensor domain: periplasmic binding domain that sense external stimuli
-Transmembrane domain: Poorly understood, but critical for sending signal to kinase domain
-Kinase domain: Histidine-containing phosphotransfer domain that is autophosphorylated upon the signal from the sensor domain being transmitted.

Question 23

The response regulator is comrpised of:

Answer 23

A receiver domain: A conserved aspartate residue that is phosphorylated by the histidine kinase
A regulatory domain: can function as a DNA-binding transcriptional regulator, an enzymatic activator (CheB) or in protein-protein interactions (CheY).

Question 24

The phosphoryl group from the phospho-HK is transferred to the ______________ of the receiver domain of the response regulator.
Phosphorylation results in ________________ of the response regulator.

Answer 24

Conserved aspartate;
conformational change

Question 25

Input signals are processed by the combined action of a highly complex regulatory network of _________________, including?

Answer 25

Interaction between genes are proteins, including activation and inhibition of genes and phosphorylation and dephosphorylation of proteins.

Question 26

The phosphorelay systems controls sporulation in B. subtilis. Describe that.

Answer 26

HK (e.g. KinA) are activated by different signals (starvation, nitrogen levels) and have a signal detection domain and a kinase domain. Ultimately, the transcription factor Spo0A is phosphorylated and this then is a negative regulator of growth and an inducer of transcription of early sporulation-specific genes. Having multiple phosphorelay proteins involved provides a series of checks and balances in order to tightly regulte sporulation initiation.

Question 27

What is chemotaxis?

Answer 27

The directed motion of an organism toward environmental conditions it deems attractive and/or away from surroundings it finds repellent.

Question 28

Movement of flagellated bacteria such as E. coli can be characterized as?

Answer 28

A sequence of smooth swimming runs (CCW flagella rotation) punctuated by intermittent tumbles (CW rotation)

Question 29

Swimming cells perform chemotaxis by ?

Answer 29

Extending their runs in the direction of favorable chemical gradients using an elaborate signaling system that controls the flagellar motor switching frequency.

Question 30

In the absence of chemoeffector gradients, the cells?

Answer 30

Swim in a random walk of runs and tumbles.

Question 31

Chemoattractant ___________ tumbling probability, extending runs and allowing cells to move up an attractant gradient.

Answer 31

Decreases

Question 32

Chemotaxis signaling pathway adapts to ___________ to allow the system to function….?

Answer 32

constant stimuli ;
in a wide range of signal concentrations

Question 33

In E. coli, what controls flagellar rotation and sensory adaptation?

Answer 33

A conserved set of cytoplasmic signaling proteins
-Che proteins

Question 34

Extracellular ligands are detected by ________________ connected to a _____________ via a linker.
- A basal rate of autophosphorylation generates ________, which transfers its phosphate to two response regulators:
_______ and _______.

Answer 34

Transmembrane chemoreceptors
Cytoplasmic kinase
CheA-P
CheB & CheY

Question 35

CheyY-P binds to the ______________, reversing its default CCW rotation to CW.
________ rapidly dephosphorylates CheY-P to maintain a ____________ and terminating the _____________ response.

Answer 35

Flagellar motor
CheZ (phosphatase)
fast response to signals
tumble

Question 36

Chemoattractant binding ________ CheA kinase activity, decreasing tumbling probability and extending runs and allowing cells to move up an attractant gradient.

Answer 36

Represses

Question 37

CheB-P does what?

Answer 37

CheB-P governs the adaptation response that returns the chemoreceptors to their presignaling state by removing methyl groups added to the receptors by the methyltransferase, CheR, so that the receptors remain responsive to changes in the chemoeffector concentration.

Question 38

E. Coli senses chemical gradients in what kind of fashion? How?

Answer 38

In temporal fashion by comparing current concentrations to those encountered over the past few seconds of travel.

Question 39

E. Coli has four transmembrane chemoreceptors, known as?

Answer 39

methyl-accepting chemotaxis proteins (MCPs) and have periplasmic ligand binding sites and conserved cytoplasmic signaling domains.

Question 40

Methyl-accepting chemotaxis proteins (MCPs) record?

Answer 40

The cell’s recent chemical past in the form of reversible methylation of specific glutamate residues in the cytoplasmic signaling domain

Question 41

How are methyl groups transferred to the specific glutamate residues of the MCPs?
How are methyl groups removed?

Answer 41

By a constitutively active methyltransferase, CheR.
Removed by the methylesterase, CheB, and released as methanol.

Question 42

_______ and _____ regulate MCP methylation state.
Upon phosphorylation by _______, _______ removes methyl groups from MCP resulting in ___________.

Answer 42

CheB ; CheR
CheA, CheB
sensory adaptation

Question 43

CheR is a _________ that acts at a constant rate, and is no modulated by ____________.
Why is demethylation required?

Answer 43

methyltransferase ; environmental stimuli
In order to avoid desensitization of MCPs.