31 Gene regulation in bacteria Lecture Outline …dstratto/bcor011_handouts/31_operons.pdf6 •The...
Transcript of 31 Gene regulation in bacteria Lecture Outline …dstratto/bcor011_handouts/31_operons.pdf6 •The...
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31 Gene regulation in bacteria Lecture Outline 11/18/05• Finish up from last time:
• Transposable elements (“jumping genes”)
• Gene Regulation in Bacteria– Transcriptional control– Cells adjust to their environment by turning genes
on and off• The operon concept
– Repressors, Inducers, Operators, Promoters• Repressible operons (e.g. trp)• Inducible operons (e.g. lac)
Transposable elements• Normal and ubiquitous
– Prokaryotes-• Genes transpose to/from cell’s
chromosome, plasmid, or a phagechromosome.
– Eukaryotes-• Genes transpose to/from same or a
different chromosome.
• Cause genetic changes– Chromosome breaks– Duplications– Knock-out genes
I’ll talk about 2 kinds:
• Insertion sequences• Ac/Ds elements in corn
• A third major class: Retrotransposons– Uses RNA intermediate and reverse transcriptase– Most Important class in mammalian genomes
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Insertion sequence (IS) elements:
• Simplest type of transposable element– Found in bacterial chromosomes and plasmids.– Encode only genes for mobilization and insertion.
Inverted terminalrepeats
Integration of an Insertion Element
Don’t worryabout thedetails, just theconcept
Staggered cut at target site
Insert IS element
Fill in the gaps
IS element carriestransposase gene
Transposaserecognizes terminalrepeats
TransposonsHave additional genes, such as those for antibiotic
resistance• (examples Tn3 (ampicillin), Tn10 (tetracycline)
Figure 18.19b
Inverted repeats Transposase gene
Insertion sequence
Insertion sequence
Antibioticresistance gene
Transposon
5′
3′
5′
3′
Barbara McClintock’s discovery oftransposons in corn:
•Kernel color alleles/traitswere “unstable”.
•McClintock concludedtransposon called “Ds”inserted into the “C” genefor colored kernels
Nobel prize, 1983
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Transposon effects on corn kernel color.
Ac activatesDs
Two transposable elementsin different sites Normal gene for
purple kernels
Ds elementinserts into colorgene andinactivates it
Ac can make transposaseDs can move, but lacks enzyme
One method forConservative Transposition
“Cut and Paste”Transposable element is cut out bytransposase and inserts in anotherlocation.
No increase in the number oftransposable elements- just achange in position
From Griffiths, Intro to Genetic Analysis
One method for replicativetransposition
From Griffiths, Intro to Genetic Analysis
Gene regulation in bacteria
But ALL organisms must adjust to changes intheir environment and all have evolvednumerous control mechanisms.
E.coli bacteria eatwhatever we eat!
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Regulation of metabolism occurs attwo levels:
– Adjusting the activity of metabolic enzymesalready present
– Regulating the genes encoding the metabolicenzymes
Figure 18.20a, b
(a) Regulation of enzyme activity
Enzyme 1
Enzyme 2
Enzyme 3
Enzyme 4
Enzyme 5
Regulationof geneexpression
Feedbackinhibition
Tryptophan
Precursor
(b) Regulation of enzyme production
Gene 2
Gene 1
Gene 3
Gene 4
Gene 5
–
–
Types of Regulated Genes
• Constitutive genes are always expressed– Tend to be vital for basic cell functions (often called
“housekeeping genes”)
• Inducible genes are normally off, but can be turnedon when substrate is present
• Common for catabolic enzymes (i.e. for the utilization ofparticular resources)
• Repressible genes are normally on, but can beturned off when the end product is abundant
• Common for anabolic enzymes
In bacteria, genes are often clusteredinto operons
Operons have:1. Several genes for metabolic enzymes2. One promoter3. An operator, or control site
(“on-off” switch)
4. A separate gene that makes a repressor oractivator protein that binds to the operator
R O 1 2 3PP
The trp Operon
5 genes: E, D, C, B, A
Same order as enzymes for trp synthesis
Controlled by a singlepromoter and operator
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More Terminology
• Repressors and Activators are proteins that bind toDNA and control transcription.
• Co-repressors and Inducers: small “effector”molecules that bind to repressors or activators
Genes of operon
Protein
Operator
Polypeptides that make upenzymes for tryptophan synthesis
Regulatorygene
RNA polymerase
Promoter
trp operon
5′
3′mRNA
trpDtrpE trpC trpB trpAtrpRDNA
mRNA
E D C B A
The trp operon: regulated synthesisof repressible enzymes
Figure 18.21a
5′
Tryptophan absent -> repressor inactive -> operon “on”
DNA
mRNA
ProteinTryptophan
(corepressor)
Active repressor
No RNA made
Tryptophan present -> repressor active -> operon “off”.Figure 18.21b
Active repressor canbind to operator andblock transcription
Tryptophan changes the shape ofthe repressor protein so it can bindDNA
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• The lac operon: regulated synthesis ofinducible enzymes
Figure 18.22a
DNA
mRNA
ProteinActiverepressor
RNApolymerase
NoRNAmade
lacZlacl
Regulatorygene
Operator
Promoter
Lactose absent, repressor active, operon off. The lac repressor is innately active, and inthe absence of lactose it switches off the operon by binding to the operator.
(a)
5′
3′
mRNA 5'
DNA
mRNA
Protein
Allolactose(inducer)
Inactiverepressor
lacl lacz lacY lacA
RNApolymerase
Permease Transacetylaseβ-Galactosidase
5′
3′
(b) Lactose present, repressor inactive, operon on. Allolactose, an isomer of lactose, derepresses the operon by inactivating the repressor. In this way, the enzymes for lactose utilization are induced.
mRNA 5′
lac operon
Figure 18.22b
Positive Gene Regulation
• Both the trp and lac operons involve negativecontrol of genes– because the operons are switched off by the
active form of the repressor protein
• Some operons are also subject to positivecontrol– Via a stimulatory activator protein, such as
catabolite activator protein (CAP)
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Promoter
Lactose present, glucose scarce (cAMP level high): abundant lac mRNA synthesized.If glucose is scarce, the high level of cAMP activates CAP, and the lac operon produces large amounts of mRNA for the lactose pathway.
(a)
CAP-binding site OperatorRNApolymerasecan bindand transcribe
InactiveCAP
ActiveCAPcAMP
DNA
Inactive lacrepressor
lacl lacZ
Figure 18.23a
– In E. coli, when glucose is always the preferredfood source
– When glucose is scarce, the lac operon isactivated by the binding of the catabolite activatorprotein (CAP)
Positive Gene Regulation- CAP • When glucose is abundant,– CAP detaches from the lac operon, which
prevents RNA polymerase from binding tothe promoter
Figure 18.23b(b) Lactose present, glucose present (cAMP level low): little lac mRNA synthesized.
When glucose is present, cAMP is scarce, and CAP is unable to stimulate transcription.
Inactive lacrepressor
InactiveCAP
DNA
RNApolymerasecan’t bind
Operator
lacl lacZ
CAP-binding site
Promoter