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Regulation of gene expression
Regulatory DNA sequences and molecules

Gene expression results in a functional gene product ( either
RNA or protein)

Genes can be constitutive (always expressed, housekeeping
genes) or regulated (expressed only under certain conditions
in all cells or in a subset of cells)

The ability to appropriately express (positive regulation) or
repress (negative regulation) genes is essential in all genes

Regulation of gene expression occurs primarily at the level of
transcription in both prokaryotes and eukaryotes and is
mediated through the binding of trans-acting proteins to cisacting regulatory elements on DNA
Regulatory DNA
sequences
and molecules

A protein transcription factor
(trans acting molecule) which
regulates a gene on chromosome 6
might itself have been
transcribed from a gene on
chromosome 11

Some trans-acting factors can
negatively affect gene expression

The binding of proteins to DNA is
through structural motifs such as
leucine zipper, or helix-turn-helix
in the protein, or zinc finger
Regulation of prokaryotic gene
expression

In eukaryotes regulation also occurs through modifications to
DNA, as well as through posttranscriptional and
postranslational events

In prokaryotes such as E.coli, the coordinate regulation of
genes- whose protein products are required for a particular
metabolic pathway- is achieved through operons

Operons: groups of genes sequentially arranged on the
chromosome along with cis-acting regulatory elements that
determine their transcription single polycistronic mRNA

Prokaryotic operon genes are turned on or off as a unit,
exhibiting both positive and negative regulation

Operon operator is DNA segment that regulates the activity
of operon structural genes
Regulation of prokaryotic gene expression

If a repressor is bound to the operator RNA polymerase is
blocked and does not produce mRNA no protein’s made

If inducer is present  it binds the repressor  repressor
changes shape and does not bind operator

If operator is not bound by a repressor RNA polymerase
transcribes protein-coding genes to mRNA  protein’s made

The lac operon contains the lacZ, lacY and lacA structural genes,
the protein products of which are needed for the catabolism of
lactose, when glucose is not available as the preferred fuel

lacY gene codes for a permease that facilitates lactose movement
into cell

lacZ gene codes for -galactosidase. It hydrolyses lactose to
galactose and glucose

lacA gene codes for thiogalactoside transacetylase (unkown job)
Lac operon

The regulatory portion of the operon is upstream of the 3
structural genes. It consists of the promoter (P) region where
RNA polymerase binds, the operator (O) site and the CAP site
(catabolite activator protein) where regulatory proteins bind

If O site is empty and CAP site is bound by cAMP-CAP
complex (CAP: catabolite activator protein or cAMP regulatory
protein-CRP-)lacZ, lacY and lacA genes are expressed

Lacl gene- a regulatory gene with its own promoter-codes for
the repressor( a trans-acting factor) that binds the operator
(O) site

Lacl gene is constitutive, its gene product, repressor, is
active. Unlike inducible lacZ, lacY and lacA genes, whose
expression is co-ordinately regulated
Lac operon
When glucose is the only sugar available

Operon is repressed (turned off)
by the binding of the repressor
protein ( the product of the lacl
gene) via a helix-turn-helix motif
to the operator (O) site, thus
preventing transcription (negative
regulation)
Lac operon
When only lactose is available


Lac operon is turned on- as a small amount of lactose is
converted to allolactose (inducer)binds repressor and
changes its conformation repressor can not bind operator
Adenylyl cyclase is actively – in the glucose absencesynthesising cAMP cAMP-CAP complex binds CAP site
RNA polymerase initiates transcription at promoter site
polycistronic mRNA(3 sets of start and stop codons)  its
translation produces 3 proteins for lactose use in energy
metabolism
Lac operon
When both glucose and lactose are available

Adenylyl cyclase is deactivated by glucose (catabolite
repression)no cAMP-CAP complex forms CAP binding
site remains empty RNA polymerase can’t transcribe 3
structural genes (even though repressor is not bound to
operator (O) site)
Regulation of eukaryotic gene expression

Gene regulation is more complex in eukaryotes. Operons are
not present, but coordinate regulation of the transcription
of genes located on different chromosomes can be achieved
through the binding of trans-acting proteins to cis-acting
elements

Posttranscriptional regulation at mRNA level include
– Alternative mRNA splicing
– Control of mRNA stability
– Control of translational efficiency

Regulation at the protein level occurs by mechanisms that
modulate stability, activation or targeting of the protein
mRNA editing
Apoprotein B(apoB) is essential
component of chylomicrons
and VLDL lipoproteins
In the intestine only, C residue
in CAA codon for glutamine
is deaminated to UUAA
stop codon  shorter
protein apoB-48 (48% of
mRNA) in the intestines,
than hepatic apoB-100(full
length)
Regulation through modifications to
DNA
• Gene expression in eukaryotes is influenced by DNA availability to
transcriptional apparatus, amount of DNA and arrangement of
DNA (localised transitions between B and Z forms)
1. Access to DNA: eukaryotic DNA is complexed with histone proteins
• Euchromatin is transcriptionally active and less condensed and
differs from heterochromatin which is more condensed and
inactive
– Active chromatin contains histone proteins modified at amino
terminal ends by acetylation or phosphorylation, decreasing
their positive charge and, hence, decreasing their association
with negatively charged DNAThis chromatin remodelling
relaxes nucleosomes, allowing transcription factors access to
DNA
Regulation through modifications to DNA
– Methylation in cytosine bases
of GC-rich regions upstream of
many genes: Transcriptionally
active genes are less methylated
than their inactive counterparts. Methylation is by
methyltransferase using S-adenosylmethionine as methyl donor
2.Amount of DNA: A change in the number of a gene copies gene can
affect the amount of gene product produced.

Gene amplification- increase in copy number- in response to
chemotherapeutic drug such as methotrexate; an inhibitor of
dihydrofolate reductase (DHFR)- is required for the synthesis of
tetrahydrofolate,.
Regulation through modifications to
DNA

Gene amplification results in an increase in the number of
DHFR genes and resistance to the drug, allowing
tetrahydrofolate to be made for DNA synthesis.
3.Arrangement of DNA: allows the generation of 109-1011
different immunoglobulins from a single gene, providing the
diversity needed for recognition of an enormous number of
antigens

Immunoglobulins – eg IgG- consist of 2 light and 2 heavy
chains, with each having regions of constant and variable amino
acid sequences

Variable region is the result of somatic recombination of
segments within both light- and heavy-chain genes
4.Mobile DNA elements: Transposons (Tn): a small mobile genetic
(DNA) element that moves around the genome or to other
genomes within the same cell, usually by copying itself to a
second site but sometimes by splicing itself out of its original
site and inserting in a new location mediated by transposase,
encoded by Tn itself

In Direct transposition, transposase cuts out and inserts Tn at a
new site. In replicative transposition; Tn copies are inserted
elsewhere while original remains in place

Transposition is basis for hemophilia A, muscular destrophy, and
antibiotic resistance ( receiving plasmids carrying transposons
with antibiotic resistance genes