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					                    Surface Motility




a) Swarming - flagella        b) Twitching – Type IV pili
                                  (Retractile motility)

c) Gliding - ?                 d) Spreading - passive
                                                                  1
                                       Harshey, Ann Rev. Microbiol. 2003
                    Swarming Motility


1. Flagella-driven group motility on a surface        Swarming Movies

2. Cell-density dependent lag prior to initiation of movement




3. Swarming colony surrounded by a wet film which facilitates movement


4. Swarmer cells are ‘differentiated’- longer and more flagella

                                                                         2
   Swarmer cells are longer and have more flagella




Broth - Swimmer                  Plate - Swarmer



                                                     3
                What are the signals?


How are the signals transduced to elicit cell differentiation?




                                                          4
Flagellar dynamometer controls swarmer cell differentiation
               of V. parahaemolyticus


                McCarter et al. 1988 Cell 54: 345-351




                                                              5
Vibrio parahaemolyticus has two different types of flagella

                                                                  H+

                   Na+   High viscosity



                         Low viscosity
                                          H+-driven
 Na+-driven                               Lateral flagella
 Polar flagellum                                             Na+-driven
 Pof                                        Laf              Polar flagellum




                                                                               6
Two flagellar systems




                        Broth: Fla

                        Na+ ions




                        Agar: Laf

                        H+ ions




                                     7
Reporter system used in this work




                                    8
Induction of laf::lux in:
                    Agar




                                 Viscosity
            PVP
                        Broth




                  PVP




                                 Antibody agglutination

                         Broth




                                                          9
      Vibrio parahaemolyticus has two different types of flagella

                                                                               H+
                                 Ab that tether Pof
                                        OR
                          Na+     High viscosity
                                 Slow motor rotation

                                  Low viscosity
                                                       H+-driven
        Na+-driven                                     Lateral flagella
        Polar flagellum                                                   Na+-driven
        Pof                                              Laf              Polar flagellum




What if you destabilize the filament?                                                       10
SDS-PAGE of purified flagella in various strains




                                                   11
EM of mutants with flagellin gene defects




                                     DFlaAB




                                      FlaC




                                              12
Vibrio parahaemolyticus has two different types of flagella

                         Ab that tether Pof                           H+
                          Mot mutations
                          Che mutations
                   Na+    High viscosity



                          Low viscosity
                                              H+-driven
 Na+-driven                                   Lateral flagella
 Polar flagellum                                                 Na+-driven
 Pof                                            Laf              Polar flagellum




                                                                                   13
Effect of fla, che and mot mutations on laf::lux expression

                        1- Fla-; 2- Fla-; 3- Che-; 4- Mot-; 5- Fla-

                        6- Fla-; 7- Mot-; 8- WT; 9- Fla-; 10- Fla-

                        11- Fla-; 12- Che-; 13- Che-; 14- Che-; 15- WT




                       WT

                                                                      14
Interference with polar flagellar function signals Laf expression


              1.     Increasing viscosity


              2.     Antibodies


              3.     FlaC mutations


              4.     Mot mutations




                                                               15
              Possible Mechanisms


1.   Slow movement signals chemotaxis in reverse


2.   High viscosity affects subunit assembly:
     negative regulation of Laf assembly by FlaC


3    Flagellar torque imparts torsional force on cell body


      All of these are physical signals!


                                                             16
The sodium-driven polar flagellar motor of marine Vibrio
as the mechanosensor that regulates lateral flagellar gene
                      expression

            Kawagishi et al., Mol. Microbiol. 20: 693-699, 1999




                                                                  17
            Hypothesis


          Rotation rate of

               OR

    External force applied against

the polar flagellum controls laf expression



                                              18
Vibrio parahaemolyticus has two different types of flagella

                           Ab that tether Pof
                            Mot mutations
                                                                        H+
                         Na+ channel blockers

                   Na+      High viscosity



                            Low viscosity
                                                H+-driven
 Na+-driven                                     Lateral flagella
 Polar flagellum                                                   Na+-driven
 Pof                                              Laf              Polar flagellum




                                                                                     19
Effect of amiloride and its analogs on




                               Growth




                              laf expression




                                               20
            Effect of

Phenamil                        Viscosity




                 on

           Swimming speed   ●

           laf expression   ○
                                            21
Relationship between swimming speed and laf induction




                                                   22
                          Conclusion



 A decrease in polar flagellar rotation rate, rather than the external force
applied against the polar flagellum, can trigger swarmer cell differentiation




                                                                       23
                 How is rotation rate sensed?

Ion flux? But flux is coupled to rotation rate, so hard to separate.

Also a marine bug like Vibrio has plenty of sodium around, so decrease
in rotation should not affect total sodium flux in cell.

Local ion concentration at the polar flagellum?

Can this change the charge/conformation of an associated regulator
of gene expression?

Is assembly still an option?




                                                                       24
Vibrio parahaemolyticus has two different types of flagella
                             Ab that tether Pof
                               Mot mutations
                          Na+ channel blockers                              H+
                         i.e. slow motor rotation
                                 Low Iron
                   Na+        High viscosity



                               Low viscosity
                                                    H+-driven
 Na+-driven                                         Lateral flagella
 Polar flagellum                                                       Na+-driven
 Pof                                                  Laf              Polar flagellum




                                                                                         25
                    Surface Motility




a) Swarming - flagella        b) Twitching – Type IV pili
                                  (Retractile motility)

c) Gliding - ?                 d) Spreading - passive
Gliding Movies                                                    26
                                       Harshey, Ann Rev. Microbiol. 2003

				
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