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Regulation of Gene ExpressionRegulation of Gene Expression

Dr. Ishtiaq Ahmad KhanDr. Ishtiaq Ahmad Khan

Dr. Panjwani Center for Molecular Medicine Dr. Panjwani Center for Molecular Medicine and Drug Researchand Drug Research

Definitions• Constitutively expressed genes:

– Genes that are actively transcribed (and translated) under all experimental conditions, at essentially all developmental stages, or in virtually all cells.

• Inducible genes:– Genes that are transcribed and translated at

higher levels in response to an inducing factor

• Repressible genes:– Genes whose transcription and translation

decreases in response to a repressing signal

Definitions

• Housekeeping genes: – genes for enzymes of central metabolic

pathways (e.g. TCA cycle)– these genes are constitutively expressed– the level of gene expression may vary

Modulators of transcription• Modulators:

(1) specificity factors, (2) repressors, (3) activators

1. Specificity factors:Alter the specificity of RNA polymerase

Examples: -factors (TBPs

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Heat shock geneHousekeeping gene Heat shock promoter

Standard promoter

Modulators of transcription2. Repressors:

mediate negative gene regulationmay impede access of RNA polymerase to the

promoteractively block transcriptionbind to specific “operator” sequences (repressor

binding sites) Repressor binding is modulated by specific effectors

Coding sequence

Repressor

Operator

Promoter

Effector(e.g. endproduct)

Negative regulation (1)

Source: Lehninger pg. 1076

Repressor

EffectorExample: lac operon

RESULT:Transcription occurs when the gene is derepressed

Negative regulation (2)

Source: Lehninger pg. 1076

Repressor

Effector (= co-repressor)Example: pur-repressor in E. coli; regulates transcription of genes involved in nucleotide metabolism

Modulators of transcription3. Activators:

mediate positive gene regulation

bind to specific regulatory DNA sequences (e.g. enhancers)

enhance the RNA polymerase -promoter interaction and actively stimulate transcription

common in eukaryotes

Coding sequence

Activator

promoter

RNA pol.

Positive regulation (1)

Source: Lehninger pg. 1076

RNA polymerase

Activator

Positive regulation (2)

Source: Lehninger pg. 1076

RNA polymerase

Activator Effector

Operons– a promoter plus a set of adjacent genes whose

gene products function together. – usually contain 2 –6 genes, (up to 20 genes)– these genes are transcribed as a polycistronic

transcript.– relatively common in prokaryotes– rare in eukaryotes

The lactose (lac) operon

• Contains several elements– lacZ gene = -galactosidase– lacY gene = galactosidase permease– lacA gene = thiogalactoside transacetylase– lacI gene = lac repressor

– Pi = promoter for the lacI gene– P = promoter for lac-operon– O1 = main operator– O2 and O3 = secondary operator sites (pseudo-operators)

Pi P Z Y A I O3 O1 O2

The lac operon consists of three structural genes, and a promoter, a terminator,regulator, and an operator. The three structural genes are: lacZ, lacY, and lacA.

• lacZ encodes β-galactosidase (LacZ), an intracellular enzyme that cleaves the disaccharide lactose

into glucose and galactose.• lacY encodes β-galactoside permease (LacY),

a membrane-bound transport protein that pumps lactose into the cell.

• lacA encodes β-galactoside transacetylase (LacA), an enzyme that transfers an acetyl group from acetyl-CoA to β-galactosides.

• Only lacZ and lacY appear to be necessary for lactose catabolism.

Theodor Hanekamp © 2003

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First Level• The lacI gene coding for the repressor lies nearby the lac operon

and is always expressed (constitutive).• Hinder production of β-galactosidase in the absence of lactose. • If lactose is missing from the growth medium, the repressor binds

very tightly to a short DNA sequence called the lac operator. • The repressor binding to the operator interferes with binding of RNA

Pol to the promoter, and therefore mRNA encoding LacZ and LacY is only made at very low levels.

• When cells are grown in the presence of lactose, however, a lactose metabolite called allolactose , which is a combination of glucose and galactose, binds to the repressor, causing a change in its shape.

• Thus altered, the repressor is unable to bind to the operator, allowing RNAP to transcribe the lac genes and thereby leading to higher levels of the encoded proteins.

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Second Level• The second control mechanism is a response to glucose, which

uses the Catabolite activator protein (CAP) to greatly increase production of β-galactosidase  in the absence of glucose. 

• Cyclic adenosine monophosphate  (cAMP) is a signal molecule whose prevalence is inversely proportional to that of glucose.

• It binds to the CAP, which in turn allows the CAP to bind to the CAP binding site (a 16 bp DNA sequence upstream of the promoter),

• which assists the RNAP in binding to the DNA. In the absence of glucose, the cAMP concentration is high and binding of CAP-cAMP to the DNA significantly increases the production of β-galactosidase

• enabling the cell to hydrolyse (digest) lactose and release galactose and glucose.

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Regulation of the lac operon

Pi P Z Y A I Q3 Q1 Q2

Inducer molecules: Allolactose: - natural inducer, degradableIPTG (Isopropylthiogalactoside)- synthetic inducer, not metabolized,

lacI repressor

Pi P Z Y A I Q3 Q1 Q2

LacZ LacY LacA

Operons in eukaryotes

Dicistronic transcription units specify a messenger RNA (mRNA) encoding two separate genes that is transported to the cytoplasm and translated in that form. Presumably, internal ribosome entry sites (IRES), or some form of translational re-initiation following the stop codon, are responsible for allowing translation of the downstream gene.

In the other type, the initial transcript is processed by 3΄ end cleavage and trans-splicing to create monocistronic mRNAs that are transported to the cytoplasm and translated.

Like bacterial operons, eukaryotic operons often result in co-expression of functionally related proteins.

19Blumenthal T, BRIEFINGS IN FUNCTIONAL GENOMICS AND PROTEOMICS. VOL 3. NO 3. 199–211. NOVEMBER 2004

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Genes whose function is to specify mitochondrial proteins and those that

encode the basic machinery for gene

expression, transcription, splicing and

translation have a very strong tendency to

be transcribed in operons

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Examples in C. elegans

An operon that expresses two subunits of the

acetylcholine receptor. An operon that encodes two proteins needed for

modifying collagen, expressed only in

collagen-producing cells. An operon that co-expresses an ion channel

protein with a protein that modifies the activity of that channel;

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