The principle of gene
control
Francois Jacob and Jacques Monod--principles of gene
control|(1961)
F.Jacob and J.Monod publish “Genetic regulatory
mechanisms in the synthesis of proteins.” A paper in
which the theory of the operon is developed. They
proposed their now-classical operon model in1961.
The core of the model was that the level of proteins in
cells was controlled at a genetic level. The theory also
predicted the existence of mRNA-an unstable
intermediate between the genome and the expressed
protein.
Before this work the prevailing model was called the
instruction hypothesis that stated that all proteins
were present in a cell, but that in the absence of an
inducer they were not properly folded and were inactive.
Jacob and Monod were awarded the Nobel Prize in
1965 for their work on characterising genes involved in
lactose utilization
-The order in the lac operon : promoter-operator-lacZ-lacY-lacA,and the
regulatory gene lacI is located close to the structural genes, just upstream
of the promoter.
-The lacI gene has its own promoter and terminator.
-The promoter for the lacI is a weak promoter, so relaticely fedw
repressor molecules are present in the cell.
-The lac repressor protein encoded by lacI is made contitutively but its
ability to bind to the operator is affected by the presence of inducer.
The lacI gene encodes the lac repressor.
The lac repressor is a sequence-specific DNA-binding protein
that Negatively regulates expression of the lac operon
structural genes.
In genetic
terminology:
The lac repressor is a
trans-acting element
trans = across
The operator is a cisacting element
cis=near
negative control
 -RNA polymerase
molecules bind to the
repressor gene(lacI+)
promoter and
transcibe the lacI
gene
 -translation produces
a polypeptides
consisting of 360
amion acids. Four of
these polypeptide
associate together to
form the functional
repressor protein.
-the functional repressor binds to the operator(lacO+). Whin the repressor is
bound to the operator, RNA polmerase can bind to the operon’s promoter but is
blocked from initiating transcription of the protein-coding genes.
The lac operon is said to be under negative control since the binding of the
repressor at the operator site blocks transciption of the structural genes.
The lactose (lac) operon of Escherichia coli -negative regulation
1. + Inducer
Induced
Repressor cannot bind
(lactose/
allolactose)
p
t
lacI
transcripts
2. No Inducer
p
o
Repressor
p
lacZ
-galactosidase
Repressor binds to laco
lacI
transcript
t
lacZ
o
lacY
Permease
lacA
Acetylase
Repressed
lacY
t
lacA
allosteric shift
-when wild –type
E.coli grows in the
presence of lactose
as the sole carbon
source, some of the
lactose transported
into the cell is
converted by existing
molecules of b galactosidase into a
metabolite of lactose
called allolactose.
- The lac repressor protein also has a recongnition site for allolactose .
-when allolactose binds to the repressor, it changes the shape of the
repressor—this is called an allosteric shift.
Repressor-inducer binding is in equilibrium
In the absence of lactose
there are only 1-5
molecules of each of the
lacZYA proteins in the
cell. Following
induction, up to 5000
molecules of galactosidase can
accumulate within
minutes.
As lactose is cleaved and used as a carbon source for growth the
lactose levels drop. Eventually free repressor molecules will
accumulate once more and the lac operon will be switched off.
Jacob and Monod used Genetic Analysis of partial
diploids to unravel the role of genes in the lac
operon.
Mutations were isolated at four loci: The genes lacZ, lacY, lacI and
in the operator lacO.
All loci were shown to be tightly linked on the E.coli chromosome
by genetic linkage analysis.
Each mutant had unique properties and behaved differently.
Analysis was achieved using Hfr strains to create partial diploids
where the new strain had two copies of the lac region.
Important mutation on negative
control
 Different gene mutation will have different
effects on the negative control.They will affect
the transcription of protein-coding genes(lacZ,
lacY and lacA).And thus affect the on and off
of the genes.
 The most important gene mutations in
negative control are:
---the lacO gene mutation(lacOc)
---the lacI gene mutation(lacI-)
F’ lacO+ lacZ - lacY+
lacOc lacZ+ lacY- (both gene sets have a normal promoter, and the
lacA gene is omitted because it is not important to our discussion)
 Results: b -
galactosidase is
synthesized in the
absence of inducer,
but permease is
not. Only when
lactose is added to
the culture and the
alllolactose
inducer is
produced does
permease syntheis
occur.
Explanations:th
e base pair
alterations of the
operator DNA
sequence make it
unrecognizable by
the repressor
protein. Since the
repressor cannot
bind, the structural
genes physically
linked to the laOc
mutation become
constitutively
expressed.
Effects of lacI gene mutations



(a) the effecst of
lacI- mutation on
lac operon
expression in a
haploid
(1)the lacI mutations
map within the
repressor structural
gene and result in
amino acid
changes in the
repressor.
(2)the repressor’s
shape is changed,
and it cannot now
recognize and bind
to the operator.
(3)As a consequence,
transcription cannot
be prevented, even
in the absence
oflactose, and
there is constitutive
expression of the lac
operon.
(b) the effects of lacI- mutation on lac operon
expression in diploid

The dominance of the
lacI+ (wild-type) gene
over lacI- mutants is
illustrated for the partial
diploid describe eariler,

lacI+ lacO+ lacZ lacY+

lacI - lacO+ lacZ+
lacY- (both gene sets
have normal operators
and normal promoters,
and the lacA gene is
omitted because it is not
important to our
discussion)

Results: In the absence of inducer,no b -galactosidaseor permease was
produce;both were synthesized in the presence of inducer. The expression of both
genes was inducible
Explanations:
 In the absence of the inducer
 The defective lacI repressor is unable to bind to either normal
operator (lacO+) in the cell. But sufficient normal repressors,
produced from the lacI+ gene, are present which bind to the two
operators and block transcription of both operons.
 (2) In the presence of the inducer
 The wild-type repressors are inactivated, so both operons are
transcribed. One produces a defective b -galactosidase and a
normal permease, while the other produces a normal b galactosidase and a defective permease; between them, active
b -galactosidase and permease are produced.
 Thus, in lacI+/lacI- partial diploids, both operons present in the
cell are under inducible control.
Other classes of lacI gene
mutation
---lacIs (superrepressor) mutatants
---lacI-d (dominance) mutants
---lacIQ (quantity) and lacISQ (superquantity)
mutants
Other classes of lacI gene mutation
lacIs (superrepressor) mutatants
In partial diploids with a lacI+/ lacIs genotype, the lacIs allele is trans- Explanations:
dominant, affecting both operon copies.The lacIs mutants, shows no (1) The mutant repressor gene
produces a superrepressor
production of lac enzymes in the presence or absence of lactose.
protein that can bind to the
operator, but cannnot
recognize the inducer
allolactose.
(2)The mutant superrepressor
bind to the operators
regardless of wherether the
inducer molecule is present
or absent, and transcription
of the operons can never
occur even in the presence
of the inducer.
(3)The presence of normal
repressors in the cell has
no effect on this situation,
since once a lacIs
repressor is on the
operator, the repressor
cannot be induced to fall
off.
Functions of the repressor protein
To conclude, specifically, the repressor is
involved in three recognition interactions, any
one of which can be affected by mutation:
i)binding of the repressor to the operator region
ii)binding of the inducer to the repressor
iii)binding of individual repressor polypeptides
to each other to form the active repressor
tetramer.
Further investigation –positive control of the lac
operon
-Several
years after Jacob and Monod proposed their
operon model, researchers also found a positive control
system that regulates the lac operon other than the
negative control
-The system function to turn on the expression of the
operon and ensure that the lac operon will be expressed
only if lactose is the sole carbon source but not if glucose
is present as well.