chapter 12 part 3 Flashcards
heat stress
45 degrees compared to 37 degrees
what do E. coli use as a result of heat stress
alternative sigma factors
what do alternative sigma factors do
activate expression of specialized heat stress response genes
- change promoter-recognition ability of the RNA polymerase core enzyme
heat shock genes
encode proteins that protect cells from certain types of heat-induced damage
at a high temperature, what sigma factor is unstable
70
what alternative sigma factor is used in high temp
32
what gene encodes sigma factor 32
rpoH
what does sigma factor 32 recognize
G-C rich sequences at the -10 position instead of the A-T rich Pribnow box
what is the promoter for rpoH recognized by
sigma factor 70 when temperature is elevated or sigma factor 24
what does high temperature also change?
chaperone proteins
chaperone proteins at normal temperature
- bind to small amount of sigma factor 32 to inhibit it from forming holoenzyme RNA pol
- targets it for degradation by proteasome
chaperone proteins at high temperature
- chaperones release sigma factor 32, leaving it free to join RNA pol
- chaperones redirected to heat-damaged cellular proteins
in bacteria, _________ regulation isn’t as common as __________ regulation
translational, transcriptional
different types of translational regulation
- translation repressor proteins
- complementary antisense RNA
where to translation repressor proteins bind
mRNAs near Shine-Dalgarno sequence
- interfere with interaction between mRNA and 16s rRNA in small subunit to prevent translation
what does complementary antisense RNA do
block mRNA translation
- creates doubled-stranded region
ex. of antisense RNA regulation
IS10 regulation
2 promoters in the IS10 insertion sequence
- Pin
- Pout
Pin
relatively weak, controls transcription of transposase
Pout
much stronger
where is the Pout promoter embedded
transposes gene
what does the Pout promoter direct transcription of
antisense RNA complimentary to part of the transposase mRNA
what happens when most of the transposase mRNA is bound to antisense RNA
very little transposase is made, transposition relatively rare
riboswitches
when an mRNA binds a small regulatory molecule
3 functions of riboswitches
- regulates transcription
- affects translation
- alters mRNA stability
a riboswitch regulates the transcription of which operon
thiamin (thi) operon
what does the thiamin operon code for
proteins of the biosynthetic pathway for thiamin pyrophosphate (TPP) production
ex. of how riboswitch alters transcritpion
riboswitch mRNA in Bacillus subtilis
low TPP concentration in Bacillus subtilis
- TPP amounts too low for riboswitch binding
- leads to antitermination stem-loop formation
- transcription of operon genes
high TPP concentration in Bacillus subtilis
- TPP binds to riboswitch
- generates termination stem-loop followed by poly-U sequence
- prevents transcription
ex. of altering translation with riboswitches
riboswitch mRNA in E. coli
what operon produces TPP in E. coli
thiMD operon
low TPP concentration in E. coli
- 5’ UTR of mrNA forms secondary structure that contains Shine-Dalgarno antisequestor stem-loop
- Shine-Dalgarno sequence binds to 16s rRNA in small ribosomal subunit so start codon may initiate translation of operon mRNA
high TPP concentration in E. coli
- TPP binds to riboswitch
- forms mRNA stem-loop that prevents Shine-Dalagarno sequence and start codon from initiating translation
- prevents translation
ex. of riboswitch troll of mRNA stability
gene glmS in B. subtilus
what does glmS code for
glutamine:fructose-6-phosphate amidotransferase
what is glutamine:fructose-6-phosphate amidotransferase involved in
production of sugar GlcN6P
when concentrations of GlcN6P are low
glmS is expressed
when concentrations of GlcN6P are high
GlcN6P binds to the riboswitch sequence in the glmS mrNA and induces mRNA cleavage and preventing translation
for successful phage infection, genetic regulatory switches must be controlled by what?
phage gene expression to redirect action of host genes
what phage is usually seen in phage infection
lambda phage
lambda phage
temperate phage that can undergo the lytic or lysogenic cycle
description of the lambda genome
composed of about 48 kb of linear double-stranded DNA, encoding nearly 60 genes
what does the lambda genome do once inside the host cell
circularizes
how does the lambda genome circularize
by joining of 2 single-stranded cohesive (cos) ends of about 12 nucleotides in length
the lambda genome is organized as a series of ________
operons
when does the expression of some operson begin in the lambda genome
immediately after circularization
3 types of lambda genes
- immediate early genes
- delayed early genes
- late genes
what lambda proteins are made first
N and cro
the products of the immediate early genes compete for control of what?
genetic switch - can be either lytic or lysogenic
where does transcription of the circularized lambda genome begin
2 promoters: PR, PL
PR promoter
- rightward transcription of early genes
- lytic cycle
- begins with Cro protein (repressor)
PL promoter
- leftward transcription of genes
- lysogeny
- transcribes N protein (antiterminator)
how is N an antiterminator
blocks transcription termination by allowing expression of delayed early and late genes
how does the regulation of lysogen begin
with protein N binding to 3 transcription terminating DNA sequences
3 transcription terminating DNA sequences
tL, tR1, tR2
tL unbound
blocks leftward transcription beyond N
tR1 and tR2 unbound
prevent rightward transcription beyond cro and 3 other early genes
what proteins are produced by leftward transcription
integrase and cIII protein
what protein is produced by rightward transcription
cII
how is cII stabilized
by binding to cIII
what does the cII/cIII complex bind
promoter PRE to initiate leftward transcription of cI gene
what is the cI protein also called
lambda repressor protein
what does the cII/cIII complex also bind to
PL to produce additional integrase protein
the genetic switch for lysis/lysogeny depends on a balance between what?
cro protein and lambda repressor (cI product)
both cro and lambda repressor have an affinity for what?
operator sequences: OR1, OR2, OR3
where are OR1, OR2, OR3
between PRM and PR
entry into the lytic cycle requires transcription of what?
late genes, regulated by late promoters and late operators
where are the late genes located
to the right of PR
what do the three operator sequences contain
17-bp target sequence for binding of either cro or lambda repressor
order of dimerized cro protein binding to operator
OR3, OR2, and OR1
what happens when the pro protein is bound at the operator sequence
blocks access of RNA polymerase at PRM, preventing production of lambda repressor (cI)
what does PRM stand for
promoter of repressor maintenance
what does cro binding also stimulate
transcription from PR enhancing transcription of cro and other rightward genes
Q gene
encodes Q protein - positive regulator of transcription of genes rightward of PR
the late genes include those needed for what?
production of head/tail proteins and proteins needed for host cell lysis
order of how dimerized lambda repressor protein binds to operator
OR1, OR2, and OR3
how does the lambda repressor function on operator
as dimer and spans 17 bap of DNA at each operator sequence
what does the lambda repressor protein bound at the operator sequences initiate
positive control at PRM - leads to continuous transcription of cI, maintains repression
- also activates integrase
a large number of co-infecting phages favors what?
lysogeny
- prevents wiping out all the host cells in a population
DNA damage to the host like UV radiation can trigger entry into what?
lytic cycle
if the host cells are actively growing, what is favored
lytic - new phage can easily find new host
if host cells are growing poorly, what is favored
lysogeny - waits until ideal conditions
what is the competition between the cro and lambda repressor protein
for binding to the operator sites
lysogeny is a ____-_________ state
semi-permanent
how can lysogeny continue for an extended time
by ongoing binding of lambda repressor to operator
induction
process that brings lysogeny to an end and reinitiates lytic cycle
what is induction primarily triggered by
DNA damage
what does DNA damage activate
DNA repair
what protein is part of the DNA repair cascade and is used in induction
RecA
what kind of activity does RecA also have
protease activity - activated by UV light damage to DNA
what happens when bacterial DNA is damaged by UV light
protease activity of RecA is activated
what does the protease activity of RecA target
lambda repressor monomers and cleaves off the C terminus
what happens when RecA targets the lambda repressor monomers
operator sequences exposed - positive regulation of cI and negative regulation of cro end
what happens when lambda repressor leaves the operator
cro protein binds to the exposed operators
what happens when cro proteins bind to the exposed operators
expression of Xis
what does Xis produce
enzyme excisionase - removes lysogen from its integrated location
