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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

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31 gene regulation in bacteria l.jpg

31 Gene regulation in bacteria


Lecture outline 11 18 05 l.jpg

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 l.jpg

Transposable elements

  • Normal and ubiquitous

    • Prokaryotes-

      • Genes transpose to/from cell’s chromosome, plasmid, or a phage chromosome.

    • 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 l.jpg

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


Insertion sequence is elements l.jpg

Insertion sequence (IS) elements:

  • Simplest type of transposable element

    • Found in bacterial chromosomes and plasmids.

    • Encode only genes for mobilization and insertion.

Inverted terminal repeats


Integration of an insertion element l.jpg

IS element carries transposase gene

Integration of an Insertion Element

Transposase recognizes terminal repeats

Don’t worry about the details, just the concept

Staggered cut at target site

Insert IS element

Fill in the gaps


Transposons l.jpg

Transposons

Have additional genes, such as those for antibiotic resistance

  • (examples Tn3 (ampicillin), Tn10 (tetracycline)

Transposon

Antibioticresistance gene

Insertion sequence

Insertion sequence

5

3

5

3

Inverted repeats

Transposase gene

Figure 18.19b


Barbara mcclintock s discovery of transposons in corn l.jpg

Barbara McClintock’s discovery of transposons in corn:

  • Kernel color alleles/traits were “unstable”.

  • McClintock concluded transposon called “Ds” inserted into the “C” gene for colored kernels

Nobel prize, 1983


Transposon effects on corn kernel color l.jpg

Transposon effects on corn kernel color.

Ac can make transposase

Ds can move, but lacks enzyme

Two transposable elements in different sites

Normal gene for purple kernels

Ac activates Ds

Ds element inserts into color gene and inactivates it


One method for conservative transposition l.jpg

One method for Conservative Transposition

“Cut and Paste”

Transposable element is cut out by transposase and inserts in another location.

No increase in the number of transposable elements- just a change in position

From Griffiths, Intro to Genetic Analysis


One method for replicative transposition l.jpg

One method for replicative transposition

From Griffiths, Intro to Genetic Analysis


Gene regulation in bacteria l.jpg

Gene regulation in bacteria

E.coli bacteria eat whatever we eat!

But ALL organisms must adjust to changes in their environment and all have evolved numerous control mechanisms.


Regulation of metabolism occurs at two levels l.jpg

(a) Regulation of enzyme activity

(b) Regulation of enzyme production

Precursor

Feedback

inhibition

Enzyme 1

Gene 1

Regulation

of gene

expression

Enzyme 2

Gene 2

Gene 3

Enzyme 3

Gene 4

Enzyme 4

Gene 5

Enzyme 5

Tryptophan

Figure 18.20a, b

Regulation of metabolism occurs at two levels:

  • Adjusting the activity of metabolic enzymes already present

  • Regulating the genes encoding the metabolic enzymes


Types of regulated genes l.jpg

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 turned on when substrate is present

    • Common for catabolic enzymes (i.e. for the utilization of particular resources)

  • Repressible genes are normally on, but can be turned off when the end product is abundant

    • Common for anabolic enzymes


  • In bacteria genes are often clustered into operons l.jpg

    In bacteria, genes are often clustered into operons

    Operons have:

    • Several genes for metabolic enzymes

    • One promoter

    • An operator, or control site

      (“on-off” switch)

    • A separate gene that makes a repressor or activatorprotein that binds to the operator

    R

    P

    P

    O

    1

    2

    3


    The trp operon l.jpg

    Controlled by a single promoter and operator

    The trp Operon

    5 genes: E, D, C, B, A

    Same order as enzymes for trp synthesis


    More terminology l.jpg

    More Terminology

    • Repressors and Activators are proteins that bind to DNA and control transcription.

    • Co-repressors and Inducers: small “effector” molecules that bind to repressors or activators


    The trp operon regulated synthesis of repressible enzymes l.jpg

    trp operon

    Promoter

    RNA polymerase

    Polypeptides that make up

    enzymes for tryptophan synthesis

    The trp operon: regulated synthesis of repressible enzymes

    Regulatory

    gene

    Genes of operon

    trpR

    trpD

    trpC

    trpB

    trpE

    trpA

    DNA

    Operator

    mRNA

    3

    5

    mRNA

    5

    C

    E

    D

    B

    A

    Protein

    Figure 18.21a

    Tryptophan absent -> repressor inactive -> operon “on”


    Slide19 l.jpg

    Active repressor can bind to operator and block transcription

    DNA

    No RNA made

    mRNA

    Protein

    Active

    repressor

    Tryptophan

    (corepressor)

    Tryptophan present -> repressor active -> operon “off”.

    Figure 18.21b


    Slide20 l.jpg

    Tryptophan changes the shape of the repressor protein so it can bind DNA


    Slide21 l.jpg

    Promoter

    Regulatorygene

    Operator

    DNA

    lacl

    lacZ

    NoRNAmade

    3

    RNApolymerase

    mRNA

    5

    Activerepressor

    Protein

    (a)

    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.

    Figure 18.22a

    • The lac operon: regulated synthesis of inducible enzymes


    Slide22 l.jpg

    lac operon

    DNA

    lacl

    lacz

    lacY

    lacA

    RNApolymerase

    3

    mRNA 5

    mRNA 5'

    mRNA

    5

    -Galactosidase

    Permease

    Transacetylase

    Protein

    Inactiverepressor

    Allolactose(inducer)

    (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.

    Figure 18.22b


    Positive gene regulation l.jpg

    Positive Gene Regulation

    • Both the trp and lac operons involve negative control of genes

      • because the operons are switched off by the active form of the repressor protein

    • Some operons are also subject to positive control

      • Via a stimulatory activator protein, such as catabolite activator protein (CAP)


    Positive gene regulation cap l.jpg

    Operator

    RNA

    polymerase

    can bindand transcribe

    Promoter

    DNA

    lacl

    lacZ

    CAP-binding site

    ActiveCAP

    cAMP

    Inactive lac

    repressor

    InactiveCAP

    (a)

    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.

    Figure 18.23a

    Positive Gene Regulation- CAP

    • In E. coli, when glucose is always the preferred food source

    • When glucose is scarce, the lac operon is activated by the binding of the catabolite activator protein (CAP)


    Slide26 l.jpg

    Promoter

    Operator

    DNA

    lacl

    lacZ

    CAP-binding site

    RNA

    polymerase

    can’t bind

    InactiveCAP

    Inactive lac

    repressor

    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.

    (b)

    Figure 18.23b

    • When glucose is abundant,

      • CAP detaches from the lac operon, which prevents RNA polymerase from binding to the promoter


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