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MBB164 > Barnes - Gene expression > Flashcards

Barnes - Gene expression Flashcards

1
Q

why do prokaryotes regulate genes?

A

efficiency - doesnt waste energy and resources
avoids chaos
adaption - cells adapt to env

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2
Q

name the 2 types of gene expression and explain them

A

constitutive: constant expression
facultative: selective expression

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3
Q

define fine and coarse control

A

fine = instant response/alterations of critical enzymes eg not making them or destroying them (achieved by covalent modification/binding of ligands)
coarse = delayed response (long-term changes +ve/-ve gene regulation
slow and economical - total cellular population of enzymes changed

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4
Q

describe methods of reversible and irreversible fine control

A

irreversible: altering enzyme activity
reversible: a/a modification eg phosphorylation
ligand binding - allosterism/feedback inhibition

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5
Q

does coarse control act on transcription, translation or both in prokaryotes?

A

both, because they are coupled

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6
Q

name 2 regulatory proteins and state the difference in control method and where they bind

A 
Repressor
- Negative control
- Binds operator
Activator
- Positive control
- Binds control elements eg regulatory sites near promoter or enhancer regions
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7
Q

state the promoter region place in e. coli

A

35 and 10 NTPs upstream transc start site in E. coli

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8
Q

describe the relationship between the consensus sequence and the promoter sequence and explain how this is similar to the shine dalgarno sequence (SD seq) in translation

A

Closer to consensus seq the promoter is, the stronger the transcription machinery binds to the DNA
Closer to SD sequence = increased translation (coarse control)

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9
Q

what property of mRNA means genes can be switched on/off quickly?

A

mRNA is metabolically unstable in bacteria and has a short 1/2 life - can respond to cellular changes more quickly

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10
Q

what is the SD (shine dalgarno) sequence

A

UAAGGAGG

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11
Q

define polycistronic transcription

A

the arrangement of several protein encoding genes needed in similar circumstances that can be transcribed and translated together

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12
Q

how does e. coli use glucose?

A

glucose –> pyruvate –> energy

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13
Q

name the 3 structural genes in e. coli’s lac operon and describe what they do

A

LacZ: beta-galactosidase (lactose to glucose and galactose as well as lactose –> allolactose)
LacY: lactose permease (transports lactose into the cell)
LacA: thiogalactoside transacetylase (removes thiogalactosides - maybe involved in detox?)

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14
Q

describe the negative control mechanism in the lac operon

A

lacI gene expressed –> LacI repressor protein transc and transl (tetramer)
LacI protein binds to operator and upstream sequence –> loop –> no RNA pol activity
incomplete repression of LacZ allows some lactose –> allolactose
when lactose is present allolactose binds LacI (undergoes allosteric change, cannot bind operator, RNA pol transcribes

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15
Q

describe the positive control mechanism in the lac operon

A 
adenylate cyclase (AC) activity inhibited by glucose transport
when glucose conc high = AC activity low = low cAMP.
CRP forms a homodimer and bind to CAP/CRP site
cAMP + CRP + DNA + RNA pol = transc
therefore when glucose is present there is low cAMP and CRP cant bind to the CRP binding region - no transcription of the downstream structural genes
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16
Q

what is IPTG? why is it used?

A

IPTG = allolactose substitute (not broken down by beta-galactosidase)
used in research to produce a protein of interest

17
Q

state which molecules are involved in the positive and negative control mechanisms in the ara operon

A

positive control:
by CRP - binds CRP binding region = transc
by arabinose - binds AraC dimer protein which binds I1 and I2 = trancs
negative control:
by arabinose - when not present AraC dimer protein binds O2 and I1

18
Q

what do clear and cloudy plaques on a lawn of E. coli indicate about the lifecycle bacteriophage lambda?

A

clear plaque: lytic - clear because the phage has lysed the cells (they’re empty)
cloudy: lysogenic - E. coli cells havent been lysed

19
Q

name the 3 lytic cycle regulators in bacteriophage λ and state what they do

A

cro: binds to DNA to control transcription
N: antitermination factor - allows transcription to carry on
Q: antitermination factor - allows transcription to carry on

20
Q

name the 3 lysogenic cycle regulators in bacteriophage λ and state what they do

A

CI: binds DNA to control transcription (at OR and OL)
CII: binds DNA to control lysogenic establishment (binds Pi and Pre)
CIII: inhibits FtsH activity

21
Q

what do genes O and P do?

A

carry out replication of the viral DNA

22
Q

how does the binding of proteins to the operator regions prevent transcription from the promoters?

A

the operator regions overlap with the transcriptional start sequences therefore a protein binding to the operator can prevent transcription

23
Q

which region in the operators does Cro have the highest affinity for? what does this mean for transcription at the promoters?

A

Cro binds with highest affinity at region 3 of OR and OL
At OR binding of Cro to region 3 blocks transc from PRM (no CI) but not from PR (Cro expression)
At OL binding of Cro to region 3 doesnt affect transcription from PL (expression of N)

24
Q

how does N work as an antitermination factor?

A

N allows expression off distal genes from PR and PL. N binding to nascent mRNA removes the secondary structures that would stop O, P and Q being transcribed

25
Q

how does Q work as an antitermination factor?

A

Q allows expression of distal genes from PR’ by binding to nascent mRNA and removing secondary structures that inhibit the transc of the structural genes

26
Q

which region in the operators does CI have the highest affinity for? what does this mean for transcription at the promoters?

A

CI binds with the highest affinity to regions 1 and 2 in the operators
at OR binding of CI to regions 1&2 blocks transc from PR (no Cro) but not from PRM (expression CI)
At OL binding of CI to regions 1&2 blocks transcription from PL (no N or CIII)

27
Q

in what 3 cases will a lysogenic prophage enter the lytic cycle?

A

1) 1/1000 spontaneous activation of prophage ‘basal induction’
2) DNA damage eg UV light in a lab
3) extreme starvation

28
Q

describe in brief how a lysogenic prophage enters the lytic cycle

A

1) starvation/DNA damage
2) protease breaks down CI
3) dissociation of CI from OR and OL
4) transcription from PL and PR
5) expression of the lytic genes

29
Q

how does a phage ‘‘decide’’ to enter lysis or lysogeny upon entering a new bacterial cell?

A 
lysis - Cro expression if:
low multiplicity of infection (MOI)
high glucose conc/rich media
severe DNA damage
lysogeny - CI expression if:
high MOI
poor media/low glucose conc
30
Q

define multiplicity of infection (MOI)

A

the ratio of viruses per bacterial cell
eg MOI = 2: 2 viruses per 1 bacterial cell
MOI = 0.5: 1 virus per 2 bacterial cells

31
Q

name and describe the 2 phases of gene expression upon entry into a new bacterial cell

A

early transcription:
nothing bound to the operators - only Cro and N expressed
delayed-early transcription:
both lytic (Cro and N) and lysogenic (CII) proteins are expressed from PL and PR

32
Q

describe how lysis is established when phage λ has entered a new bacterial cell

A

Cro, N, CII expressed in delayed-early transcription
Low MOI/high glucose conc = low cAMP = FtsH not repressed therefore degrades CII
Low CII levels means there is no expression from PRE of CI
therefore Cro can bind OR and OL without competition from CI - silencing of PRM

33
Q

describe how lysogeny is established when phage λ enters a new bacterial cell

A

Cro, N and CII expressed in delayed-early transcription
High MOI/low glucose = high cAMP = FtsH repressed = no degradation of CII
CII accumulates above threshold value
CII binds PRE
CI expressed from PRE
CI binds Or and OL before Cro - silencing of PR and PL

34
Q

describe the maintenance of lysis in phage λ

A

Cro bound to OR and OL from establishment of lysis - PR and PL expressed
CIII expressed PL = inhibition of FtsH
N antitermination of PL
Q antitermination of PR’
expression of lytic genes and structural genes

35
Q

describe the maintenance of lysogeny in phage λ

A

CI bound to OR and OL from lysogeny establishment - silencing of PR and PL and expression PRM
CI expressed from PRM

MBB164 (9 decks)

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