Chapter 7

Chapter 7: Microbial Regulatory Systems

7.1 DNA-Binding Proteins

  • Gene Arrangement:

    • Distinction between Bacterial/Archaeal vs. Eukaryotic:

    • Bacteria and Archaea lack introns.

    • Genes can be organized in operons:

      • Definition: Operons consist of two or more genes transcribed under a single promoter.

      • The promoter region: Single regulatory site, positioned upstream where RNA polymerase initiates transcription.

  • Characteristics of Bacterial and Archaeal Promoters:

    • Defined by specific nucleotide sequences recognized and bound by DNA-binding proteins.

    • Role: Facilitate RNA polymerase binding and promote transcription.

  • Small molecules influence binding of regulatory proteins, which turns transcription on/off

    • Major site of binding is on major groove of DNA

  • Interaction of proteins

    • Inverted repeats are nucleotide sequence followed downstream by inverted complement

    • Homodimeric-two identical peptides that bind to inverted repeats

7.2 Transcription Factors and Effectors

  • Effect of Transcription on mRNA and Protein Production:

    • Increased transcription leads to higher mRNA levels, resulting in more protein synthesis.

  • Transcription Factors:

    • Function: Proteins that regulate transcription rates through specific DNA binding.

    • Activator proteins (Figure 7.4a):

      • Turn on transcription by binding to DNA and recruiting RNA polymerase or sigma factor.

    • Repressor proteins:

      • Turn off gene expression by binding to operator regions of DNA downstream from the promoter.

  • Structure of Transcription Regulatory Parts:

    • Part: Repressor/Activator + binding site

    • Modulator: Effector (activator or co-repressor), Environmental signal.

    • Circuitry: Combination of binding sites and parts

  • Effectors:

    • Definition: Small molecules influencing activator and repressor binding.

    • Commonly include cellular metabolites (substances, products) or structural analogs.

  • Allosteric Proteins:

    • Definition: Proteins whose conformation changes upon effector molecule binding.

    • Role in Transcription:

    • Altered conformation determines the ability of transcription factors to bind DNA.

    • Inducers: Promote transcription

    • Corepressors: Inhibit transcription.

7.3 Repression and Activation

  • Enzyme Repression:

    • Process: Prevents enzyme synthesis in the presence of its product, thereby reducing synthesis.

    • Characteristic: Does not affect synthesis of other enzymes (specific effect).

    • Common in amino acid and nucleotide biosynthesis.

    • Corepressor: The final product of a biosynthetic pathway.

  • Enzyme Induction:

    • Opposite of repression; it stimulates enzyme production in response to a substrate's presence.

    • Typically regulates degradative/catabolic enzymes (e.g., lac operon).

  • Mechanisms of Repression and Derepression:

    • Repressors inhibit transcription, while corepressors bind DNA only when the effector is present.

    • Example: In arginine regulation, arginine acts as a corepressor and binds arginine repressor (ArgR)

    • Example: Lac operon demonstrated by LacI repressor action, which blocks transcription until an effector (inducer) is present.

      • Corepressor=inducer

    • Terms:

      • Negative control: Inhibition of transcription by repressors.

      • Basal transcription: Low-level transcription occurring even when fully repressed.

  • Mechanisms of Activation:

    • Positive control: Regulator protein facilitates transcription

    • Some operons require an activator protein to promote transcription.

    • Activation occurs when an activator binds to poor promoter sequences, assisting RNA polymerase binding.

    • Example: Maltose catabolism (MalT cannot bind DNA without maltose).

      • Controlled by same regulatory protein called a regulon

    • Specifically bind to activator binding site

7.4 Transcription Controls in Archaea

  • Archaeal regulation resembles bacterial control more than eukaryotic.

    • Archaea in positive and negative control

  • Regulators: Both positive and negative controls used to modulate transcription.

  • Repressors: Block RNA polymerase binding or the binding of key transcription factors (TBP and TFB).

  • Activators: Recruit TBP to promoters to enhance transcription.

7.5 Two-Component Regulatory Systems

  • Prokaryotic Response:

    • Regulation of metabolism in response to environmental changes via two-component systems.

    • Signal may be detected by signal, signal transduction

  • Two-Component Regulatory System:

    • Sensor Kinase: In cytoplasmic membrane and autophosphorylates upon detecting a signal (histidine kinase).

    • Response Regulator: In cytoplasm and DNA-binding protein in the cytoplasm that regulates transcription.

  • Feedback Loop:

    • When the response terminates, a phosphatase removes the phosphate from the response regulator to reset the system.

7.6 Regulation of Chemotaxis

  • Chemotaxis defined: Moving towards attractants and away from repellents, responding to temporal gradients.

  • Functionality: Modified two-component system drives regulation, interfacing at the flagellar level to control preexisting proteins rather than modifying transcription.

  • Signal Mechanics:

    • Methyl-accepting chemotaxis proteins (MCPs): Key sensors for detecting attractants and repellents.

    • Interaction Leads:

      • Activation of CheA (sensor kinase) leading to autophosphorylation.

      • Phosphorylated CheY regulates flagellar rotation direction (counterclockwise vs. clockwise).

  • Adaptation Mechanism: Stops the response, relying on CheB for feedback.

    • Feedback resets system

  • Importance of Che Proteins: Also involved in phototaxis (light) and aerotaxis (oxygen).

7.7 Cell-to-Cell Signaling

  • Communication Mechanism:

    • Prokaryotes utilize small extracellular molecules to communicate.

    • Quorum sensing defined: Mechanism allowing bacteria and archaea to assess population density before launching coordinated activities.

  • Mechanism:

    • Each species produces autoinducer signaling molecules that diffuse across the cell envelope.

    • Accumulation triggers transcription of specific genes through binding at activation proteins or sensor kinases.

  • Types of Autoinducers:

    • Acyl homoserine lactone (AHL) identified in Gram-negative bacteria

    • AI-2 for interspecies communication

    • Peptide autoinducers for gram-positive bacteria.

  • Application in Pathogenesis:

    • E. coli O157:H7 produces AHL AI-3, activating virulence genes

    • Staphylococcus aureus secretes peptides for immune system interference, under control of autoinducing peptide (AIP)

7.8 The lac Operon

  • Global Control:

    • Repression mechanisms that govern sugar catabolism.

  • Catabolite Repression:

    • Order of carbon source utilization: glucose first, regardless of availability of other sugars (e.g., lactose).

    • Results in diauxic growth phases

      • Two exponential growth phases

    • Best source-growth stops-resumes with 2nd source

  • Cyclic AMP and CRP Regulation:

    • High levels of cyclic AMP promote binding of CRP activator to lac promoter, enabling transcription.

      • Allosteric and binds to cAMP

        • Regulatory nucleotide derived from adenosine

        • Synthesized by adenylate cyclase

    • The presence of an inducer prevents LacI from binding, allowing transcription to occur.

7.9 Stringent and General Stress Responses

  • Stringent Response:

    • Mechanism enabling survival under nutrient deprivation, environmental stress, or in the presence of antibiotics.

    • Triggers shut down of macromolecule synthesis while activating survival pathways.

    • Governed by alarmones:
      guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp) alarmones produced by RelA and SpoT proteins.

  • General Stress Response:

    • Driven by RpoS (stationary phase sigma factor), addressing nutrient limitation and other stresses, with regulation of over 400 genes.

7.11 The Heat Shock Response

  • Definition: A global control mechanism to protect cells from protein denaturation due to heat and other stressors.

  • Heat Shock Proteins counteract damage of denatured proteins and help cells recover from stress:

    • Classes include Hsp100, Hsp90, Hsp70, Hsp60, and Hsp10, which aid in protein refolding and degradation.

  • Role of RpoH:

    • Controls expression of heat shock proteins and is regulated by DnaK levels.

    • RpoH mRNA basepairs with itself and regulates translation

7.12 Regulatory RNAs

  • Noncoding RNA (ncRNA):

    • Categories include rRNA, tRNA, and small RNA (sRNA): 40–400 nucleotides regulating gene expression.

    • Mechanism: Base pairing with target mRNAs to modulate translation rates, preventing double-stranded RNA from being translated.

7.13 Riboswitches

  • Functionality: RNA can recognize and bind to other molecules, acting as ribozymes or regulating gene expression (ribiswitches).

  • Mechanism: Metabolite binding alters mRNA structures and affects transcription or translation.

    • Aptamer region is regognition domain that binds small molecules

    • Alteration controls expression platform

7.14 Attenuation

  • Definition: Transcriptional mechanism allowing premature termination of mRNA synthesis, controlling output beyond initiation.

    • Example: Tryptophan operon regulation demonstrates this mechanism.

  • Leader is first part of mRNA structure that can fold into other structures, encourage leader peptide

    • trp-synthesis

    • No trp-no synthesis

Feedback Inhibition

  • Definition: A regulatory mechanism for temporarily shutting off reactions in a biochemical pathway.

    • End product binds to an initial pathway enzyme, halting pathway function.

    • Feedback inhibition is reversible based on changes in product levels.

      • Binding at allosteric site changes conformation