Bacterial Gene Regulation

Regulation of Bacterial Genes

• A crucial step in bacterial gene regulation occurs at transcription initiation.

Lac Operon

• A null mutation in the gene encoding Lac repressor results in constitutive expression of the lac operon.

Coupled Transcription and Translation

• Transcription and translation are coupled in bacteria, but not eukaryotes, because prokaryotes lack a nuclear membrane.

Negative Regulation and Transcription

• Negative regulators, like the Lac repressor, prevent RNA polymerase from initiating transcription by physically blocking the DNA binding site of RNA polymerase.

Positive Regulators and Transcription

• Positive regulators affect transcription by RNA polymerase in prokaryotes by interacting with RNA polymerase to increase the frequency of transcription initiation.

Allosteric Proteins

• Proteins that change conformation when bound to an effector molecule are called allosteric proteins.

Catabolic Pathway Regulation

• Catabolic pathways are regulated in response to the levels of the molecule that is to be broken down.

Anabolic Pathway Regulation

• Anabolic pathways are regulated in response to end product of the pathway.

Global Gene Regulation in Bacteria

• Alternative sigma factors are a mechanism of global gene regulation, recognizing different promoters.

Glucose and Catabolite Repression

• Glucose causes cAMP levels to decrease, which leads to decreased CRP binding, repressing the lac operon even when lactose is present.

Helix-Turn-Helix Motif

• A helix-turn-helix motif in a transcription factor binds a specific sequence in the major groove of DNA.

Tryptophan and the trp Operon

• Tryptophan, the end product of the trp operon, changes the conformation of the repressor, enabling it to bind DNA and block transcription.

Trp Operon Regulation

• Attenuation explains regulation of the trp operon in trpR– mutants suggesting the mechanism is repressor-independent

Heat-Shock Response in E. coli

• In E. coli, the heat-shock response is mediated by the synthesis of alternative sigma factors at high temperatures that activate transcription of heat-shock genes.

Gene Regulation and Operons

• Regulatory genes encode trans-acting proteins that interact with cis-acting DNA elements.

Trp Operon and Attenuation

• In the trp operon, mutating the two Trp codons in the leader sequence to stop codons leads to region 3 binding to region 4. This means transcription will proceed regardless of Trp presence.

Transcription Initiation

• Initiation of transcription by RNA polymerase involves the binding of the sigma subunit to the core enzyme.

Transcriptional Transition

• The transition from transcriptional initiation to elongation involves the release of the sigma factor.

Effector Molecules

• Allolactose and Trp are examples of effector molecules.

Operons

• A single DNA unit that enables the simultaneous regulation of more than one coding region in response to environmental changes is called an operon.

DNA Site Alterations

• Alterations to DNA sites, such as promoters and operators, can act only in cis.

Operon Theory

• Monod and Jacob proposed the operon theory.

Reporter Genes

• The lacZ coding sequence under the control of a prokaryotic promoter serves as an example of a reporter gene.

Sigma Factors and Heat-Shock

• Sigma 32 is the sigma factor that mediates a global heat-shock response in E. coli.

Tryptophan Regulation

• In the regulation of the trp operon, tryptophan acts as an effector.

Bacterial Strains

• The data for the fifteen bacterial strains suggests all strains have mutations that prevent transcription of the operon.

Transcription Factor Advantage

• It is advantageous for transcription factors to be multimeric and for their binding sites to be clustered because the strength of DNA-protein interactions is increased.

Small RNAs

• Small RNAs regulate gene expression by binding target RNAs and inhibiting translation.

Promoters for Antisense RNA

• Promoters for antisense RNA and target genes are oriented to transcribe opposite DNA strands of the same segment.

Lac Regulatory Region

• Using the lac regulatory region to control the expression of exogenous proteins in bacterial cells allows expression to be controlled by adding an inducer to the media.

Transcriptome Analysis

• Analysis of transcriptomes is useful for genetics research because it can reveal aspects of gene regulation.

E. coli and Bioluminescence

• E. coli aided by the cloning of bioluminescence genes from Vibrio fischeri, gene fragments from V. fischeri were cloned into plasmids, transformed into E. coli, and then retransformed back into V. fischeri.

V. fischeri Density Sensing

• V. fischeri senses the density of V. fischeri cells by secretion of a molecule that can be internalized and bound to a receptor in the cytoplasm.

Targeting Quorum Sensing Mechanisms

• Targeting quorum-sensing mechanisms for antibiotic development may have no selective advantage to individual cells that are resistant to quorum-sensing inhibitors.

Ribosome Binding Mutation

• A mutation in a ribosome binding site in the lac operon would likely result in reduction in the amount of LacZ, LacY, or LacA protein.

Bacterial Genotype

• Cells of genotype I+ oc Z + Y − / F' (I+ o+ ZR Y+) would be able to utilize lactose as an energy source.

Protein-DNA Interactions

• Some bands on the gel are absent in the sample with the transcriptional regulator, therefore the protein binds the DNA fragment.

Enzyme Function

• An enzyme functions in a biosynthetic pathway in bacteria. The product of the reaction regulates levels of the protein by binding an aptamer in the RNA leader sequence that alters the conformation to block the ribosome binding site.

Prokaryotic Genome

• If you identified five mRNAs that are not translated in these mutant bacteria, the five mRNAs are transcribed from five different genes located far from the deletion. This indicated the deletion prevents expression of a small RNA that regulates translation of all five genes.

Computer Analysis

• When RNA-Seq is performed before and after heat shock of bacterial cells, computer analysis would be expected to show after heat shock, an increase in the number of sequence reads of cDNAs that encode molecular chaperone proteins.