MICR 221 Lecture 12: Motility and Chemotaxis PDF

Lecture 12: Motility and Chemotaxis Feb. 3, 2025 1 Lecture Learning Outcomes After this lecture, students will be able to describe… The major components of bacterial flagella, and how they function in flagellar swimming How the direction of flagellar...

Lecture 12: Motility and Chemotaxis Feb. 3, 2025 1 Lecture Learning Outcomes After this lecture, students will be able to describe… The major components of bacterial flagella, and how they function in flagellar swimming How the direction of flagellar rotation contributes to the process of chemotaxis How MCPs control the direction of flagellar rotation through ligand binding and methylation General features of swarming motility and twitching motility 2 Bacterial Motility and Taxis Many bacteria are motile flagellum Able to move Most use flagella or pili Taxis: directed movement in response to a stimulus Positive taxis: movement towards stimulus E.g., nutrients Negative taxis: movement away from stimulus E.g., toxic substances Can detect chemical stimuli Chemotaxis: move towards or pili away from attractants and repellents, respectively 3  Flagella Long helical surface structures 20 nm diameter, up to 20 μm long Mainly used for motility in liquid environments Flagellar swimming results from rapid rotation of flagella Water is very viscous for microbes Flagella rotate 100-1000 times per second Can swim up to 100 μm/s 4 Image from: Kearns, D.B., Nat. Rev. Microbiol., 2010, 8, 634 Flagella Number and distribution vary Atrichous: no flagella Monotrichous: flagellum on one pole Lophotrichous: tuft of flagella at one or both poles Amphitrichous: single flagellum on both poles Peritrichous: flagella distributed over surface 5  Flagellum Structure Basal body Anchors flagellum to cell envelope Contains motor (responsible for rotation) Filament Long helical structure that extends from surface Rotation moves cell Hook Flexible, bent structure Transmits rotation from basal body to filament Gram-Positive Gram-Negative 6 Image from: Prescott’s Microbiology, 11th Edn  Basal Body Protein-based structure composed of central rod, several rings Most Gram-negatives have: L, P, and MS rings embedded in cell envelope C ring in cytoplasm (not shown) Export functions Basal body helps build hook, filament during flagellum assembly Motor functions Basal body rotates hook, filament 7 Image from: Tan et al., Cell, 2021, 184, 2665  Flagellar Motors Motor built from: Rotor (rod, MS ring) Bushings/bearings (P, L rings) Stator (MotA, MotB) Rotation usually powered by proton motive force Proton channel: MotA, MotB Switch (C ring) determines direction of rotation 8 Image from: Prescott’s Microbiology, 11th Edn Video from: @bradyajohnston; https://imgur.com/a/jwaRE0g  Flagellum Assembly Basal body assembles first, then hook, then filament Subunits exported up hollow core, added to distal end Filament made of thousands of flagellin proteins Assembled in helical pattern Incorporated under cap protein 9 Image from: Prescott’s Microbiology, 11th Edn  Flagella and the Immune System Flagella are good immune targets Surface-exposed Innate immune system can recognize flagellin Toll-like receptor 5 (TLR5) Binding of flagellin to TLR5 activates NF-κB Transcription factor Leads to production of pro- NF-κB activated inflammatory cytokines 10 Image from: https://www.invivogen.com/anti-htlr5-iga  Flagella and the Immune System Flagella also targeted by adaptive immune system Major antigenic structure E.g., E. coli O157:H7 H7: flagellar antigen Contain 1000s of copies of protein subunits E.g., flagellin Increases likelihood of antibody formation Antibody binding leads to phagocytosis 11 Image from: https://www.sciencephoto.com/media/440107/view/tem-of-the-bacteria-proteus-mirabilis Immune Evasion and Phase Variation Once in host, some bacteria stop producing flagella Some bacteria alternate between different flagellins Phase variation (reversible change in phenotype) E.g., Salmonella enterica has flagellins FljB and FliC Reversible inversion of promoter (recombination) 12 Chemotaxis: Random Walk If no chemical gradient, no benefit to moving in a particular direction Bacteria move in a random walk Swim forward (run) Occasionally reorient (tumble) Swim forward in new random direction (run) 13 Flagellum Rotation Direction of flagellum rotation determines whether cell is running or tumbling Monotrichous bacteria: Peritrichous bacteria: 14 Chemotaxis: Biased Random Walk In a gradient, some directions are advantageous Move in biased random walk If direction is favourable: Longer runs Fewer tumbles Tumbles more frequent if direction is unfavourable 15  Methyl-Accepting Chemotaxis Proteins How do chemical stimuli determine direction of rotation? Methyl-accepting chemotaxis proteins (MCPs) Chemoreceptors in cytoplasmic membrane Different MCPs sense different attractants, repellents 16 Image from: Prescott’s Microbiology, 11th Edn MCPs and CheW By default, flagella rotate counterclockwise Favours runs MCPs can signal to flagella to change direction MCP ligand-binding domain forms complex with ligand Ligand changes conformation of MCP signalling domain CheW is bound to MCP signalling domain Presence of ligand impacts CheW activity 17 If No Attractant Ligand is Bound to MCP… If direction is “bad”, cell wants to tumble to reorient: 1 - CheW detects no attractant bound to MCP 2 - CheW causes kinase CheA to be auto-phosphorylated 3 - CheA-P phosphorylates CheY 4 - CheY-P binds to flagellum switch, causing CW rotation 5 - Over time, CheY-P is dephosphorylated by CheZ 18 If Attractant Ligand is Bound to MCP… If direction is “good”, cell wants to keep running forward: CheW detects attractant bound to MCP CheW does not cause CheA to be auto-phosphorylated CheY not phosphorylated, doesn’t bind to flagellum switch CCW rotation is favoured 19 MCP Excitation Direction of rotation is based on input from many MCPs Ligand binding causes counterclockwise (CCW) bias More MCPs with bound ligand → less CheY-P formed Longer runs, fewer tumbles 20  MCP Adaptation ligand Remember, ligand binding to MCP: Decreases CheA autophosphorylation Decreases amount of CheY-P A few seconds after ligand binds, MCP is methylated by CheR Methyl-accepting chemotaxis protein Methylation of MCP: Increases CheA autophosphorylation Increases amount of CheY-P 21 Image from: https://doi.org/10.1038/nrm1524 MCP Adaptation Methylation offsets impact of ligand binding on flagellar rotation; removes CCW bias If ligand leaves, methylated MCP still enhances CheA activity; causes CW bias 22  Temporal Gradients How do bacteria measure concentration gradients for chemotaxis? Usually temporally, not spatially Current concentration → number of MCPs with ligands Past concentration → number of methylated MCPs Difference results from the ~2 s delay of methylation after ligand binding Allows bacteria to swim up or down gradients 23 Image from: https://doi.org/10.1371/journal.pcbi.1005966 Swimming Up a Gradient If ligand concentration is increasing: More ligands bind to MCPs More MCPs are methylated (but this is delayed) Result: CCW bias (longer runs, fewer tumbles) 24 Swimming Down a Gradient If ligand concentration is decreasing: Fewer ligands bind to MCPs Methylation may increase (due to delay), or stay the same Result: CW bias (shorter runs, more tumbles) 25 MCP Demethylation MCPs are demethylated by CheB over time Once an MCP is demethylated, ligand binding will again result in CCW bias 26 Importance of Demethylation Demethylation makes it possible to respond to gradients E.g., increasing attractant concentration normally causes CCW bias But, if demethylation did not occur, more attractant binding wouldn’t cause CCW bias: 27  Swarming Motility Flagella can also be used to move on solid media Swarming: coordinated movement of groups of bacteria (rafts) across surfaces Non-Swarming Swarming Requires multiple flagella (usually peritrichous) Also requires surfactants Reduces surface tension between cell and surface Some are made, secreted by bacteria 28 Image from: Kearns, D.B., Nat. Rev. Microbiol., 2010, 8, 634  Swarming Motility Proteus mirabilis Common cause of catheter-associated UTIs Form swarmer cells on surfaces Produce 1000s of flagella, become 20 - 50X longer Form multicellular rafts Rafts can swarm along catheter, enter urinary tract Form crystalline biofilms which can block urethral catheters 29 Image from: https://doi.org/10.1128/IAI.72.7.3941-3950.2004  Twitching Motility Surface motility using type IV pili Pilus extends, binds to surface, retracts Pulls cell toward attachment site Jerky/twitching motion Attach to inert surfaces, other cells Used to colonize new environments E.g., host colonization by pathogens 30 Image from: https://doi.org/10.1111/j.1574-6976.2011.00307.x ; Video from: https://doi.org/10.1146/annurev-micro-092611-150055  Twitching Motility Pilus extends by adding subunits to base Pilins (e.g., PilA, blue circles) After adhering to surface, pilus then rapidly retracts Pilins removed from base ~1500 per second 31 Image from: https://doi.org/10.1111/j.1574-6976.2011.00307.x https://doi.org/10.1038/s41579-019-0195-4 Reminders Bacteriology Quiz 3 closes Feb. 4, 3 PM Lectures 9 - 11 Office Hour: Tuesday, Feb. 4, 12 – 1 PM, Botterell 449 Strike-related activities could impact the MICR 221 lab. The original schedule for Labs, Lab Assignments, and Pre- Lab Quizzes can be found on the onQ Timeline page. Any changes to the lab schedule will be announced on onQ. 32