chapter 10 lecture outline molecular regulation. microbiology: an evolving science © 2009 w. w....

Chapter 10Lecture Outline

Molecular Regulation

Microbiology: An Evolving Science © 2009 W. W. Norton & Company, Inc. 2

Microbial Gene Organization

Promotor

Activator/Operator

Leader

Structural genes(transcribed)

DNA-dep. RNA polymerase binds

Helps align mRNA in ribosomes

On-Off switch

Operon•Multiple genes transcribed from one promoter•Genes are transcribed together


Regulating Gene Expression Microbes monitor two different compartments

Intracellular Concentrations of vitamins, sugars, amino acids, nucleotides Control of de novo synthesis and degrading enzymes

Extracellular What type of environment (e.g., pond water, gastrointestinal

tract, in a host cell) Adjustment of gene expression for best protection/invasion


Regulating Gene Expression Microbes must sense their environment

Receptors on cell surface Receptors must transmit information to chromosome Signals from activated receptor alter gene expression

Change transcription rateChange translation rate


Response to Conditions Outside of the Cell Sensing the Environment Two-component signal transduction

Sensor kinase protein in plasma membrane

Binds to signal Nutrient Chemical cue

Activates itself via phosphorylation Cytoplasmic response regulator

Takes phosphate from sensor Binds chromosome

Alters transcription rate of multiple genes

Or Activator


Microbial Control of Gene Expression Occurs at Multiple Levels Alteration of DNA sequences

E.g., phase variation Control of transcription

Operators = sequences on DNA / repressors = proteins binding to DNA

Activators = sequences on DNA / activators = proteins binding to DNA

Control of mRNA stability Translational control Post-translational control

Modifying protein activity by chemical modification like phosphorylation, methylation, acetylation etc.


Control of Gene Transcription Regulatory proteins bind to DNA regulatory sequences that control

gene transcription Operators: binding of regulator (repressor) down regulates expression

of target genes Activators: binding of regulator (activator) increases expression of

target genes Regulatory proteins (regulators) bind to small molecular weight

compounds (ligands) Different regulators bind to different ligands Once ligand is bound to regulator the ability of regulator to bind to

DNA is altered


Gene Regulation via Repressors Repressors (protein) bind to operators (DNA sequence) Upon binding to the operators repressors inhibit gene

transcription Two scenarios are possible:

Active repressor is removed from operator by ligand (inducer) binding

Gene was off now gene on Gene induction

Inactive repressor binds to operator after binding of ligand (co-repressor)

Gene was on now gene off Gene repression

If ligand diminishes and not available for repressor binding, repressor releases from operator

Gene was off now gene on Gene derepression


Gene Regulation via Repressors

Active Inactive

Lactose operon Tryptophan operon


The E. coli lac Operon Lactose (milk sugar) is used as nutrient

Cannot pass through plasma membrane Lactose permease allows entry Proton motive force used to move lactose inside cell

Must be converted to glucose to be digested -galactosidase converts lactose to glucose

Humans also make -galactosidase If not, person is lactose-intolerant

Inducer


The E. coli lac Operon The lacZ gene encodes -galactosidase The lacY gene encodes lactose permease

Need both proteins to digest lactose

Role unclear


The E. coli lac Operon

© 2008 W.W. Norton & Company, Inc. MICROBIOLOGY 1/e

Repressor protein LacI blocks transcription Active repressor binds to operator

Repressor responds to presence of lactose Binds inducer (allolactose) or DNA, not both Add lactose in the absence of -galactosidase

allolactose accumulates repressor falls off operator

allolactose


The E. coli lac OperonAnimation: The lac Operon

Click box to launch animation


Concept QuizWhat is the phenotype of an E. coli strain with a mutation in the lac operon operator such that Lac repressor could never bind to the operator?

a. The strain never transcribes lac genes.

b. The strain transcribes lacZ and lacY only when lactose is present.

c. The strain always transcribes lacZ and lacY, even without lactose present.


Trp Operon Cell must make the amino acid tryptophan

Requires many proteins, made from one operon When tryptophan is plentiful, cell stops synthesis

Trp repressor must bind tryptophan to bind DNA Opposite of lac repressor

Repressor + Tryptophan

Transcription repressed


Gene Regulation via Activators Activators proteins typically bind poorly to

activator DNA unless an inducer is present If inducer concentration diminishes activator

protein can no longer bind to activator DNA and gene transcription ceases


Sigma Factor Regulation Allow coordinated expression of large gene sets Normally degraded Under stress no degradation

Binding to core RNA Polymerase Sigma factor/RNA polymerase complex binds to various heat-shock genes

By controlling factor expression large gene sets can be controlled factors regulate transcription of all genes

70 initiates transcription at most genes Alternative factors used for special cases

Sporulation in B. subtilis—Bs 28

Stationary phase—38

Heat-shock and stress response—32

Flagellar synthesis—28

Control of ratio of factors determines global control of protein synthesis


Sigma Factor Regulation Temperature-sensitive mRNA structure

Allows translation of 32 only at high temperature.

Proteolysis rapidly removesfactorsRapid turnover allows more exact control

70 degraded rapidly at 42°C

Synthesis of proteins that inhibit factorsAnti-factors block activity until needed

Anti-anti- factors respond to environment


Small Regulatory RNAs Many intergenic regions encode small untranslated RNA

(sRNA) Small RNAs regulate transcription or stability of specific

mRNA nmolecules Does not require protein translation

Economical regulation Antisense RNA base-pairs to mRNA

Usually prevents translation Until removed via endonuclease

Universal method of gene control—found in all creatures


Quorum Sensing Cells work together at high cell density

V. fischeri becomes bioluminescent Many bacteria form biofilms Exoenzyme production Toxin production

Send signal chemical to other cells Chemical accumulation = high cell density

Autoinducer

Binds to sensor in cell Sensor activates transcription


Microbial Communication through Quorum Sensing

LuxI synthesizes the autoinducer homoserine lactone

Autoinducer diffuses into the medium where it accumulates. At threshold concentration AI diffuses back into the cell and binds to activator protein LuxR.


Peptides

chapter 10 lecture outline molecular regulation. microbiology: an evolving science © 2009 w. w....

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evolving science

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gene regulation

proteins binding

dna repressors

gene transcriptiontwo

dna regulatory sequences

dna activators