Molecular basis of type IV pilus-dependent motility in Myxococcus xanthus: Mechanism, regulation and nanomechanical analysis.

摘要

Myxococcus xanthus is a Gram-negative soil bacterium that exhibits complex social behavior. It forms swarming groups under vegetative conditions and aggregates into fruiting bodies under starvation. When starvation is prolonged, cells in the fruiting bodies undergo sporulation. M. xanthus moves over solid surfaces via two motility systems: adventurous (A) motility for individual cell movement, and social (S) motility for coordinated group movement. S-motility plays an essential role in the social lifestyle of M. xanthus, and this dissertation addresses the mechanism and regulation of social motility in M. xanthus.; Two extracellular structures, fibril material and type IV pilus (TFP), are known to be essential for S-motility. TFP powers social motility via attachment to a solid surface and retraction, which pulls the cells forward. The function of extracellular fibril material in S-motility remains elusive. A retraction assay was developed in this study, demonstrating that fibril material can trigger the retraction of TFP, and this retraction can be blocked with an antibody generated against the native PilA (pilin) protein. Further analysis identified amine-sugars in fibril material as the active components for this bioactivity.; The main regulator for TFP function in S-motility is the Frz signal transduction system, which controls pili pole-to-pole switching frequency. The output signal from the Frz system is unknown. Genetic approaches were taken to investigate the role of FrzE, the last component of the Frz pathway. Analysis of motility phenotypes of the resulting mutants revealed that the C-terminal domain of FrzE protein plays a pivotal role in controlling and coordinating both A- and S-motilities in M. xanthus.; In the last part of this dissertation, atomic force microscopy (AFM) was applied to image the cellular ultrastructures (e.g. TFP, fibril material), revealing their morphological details under native conditions and at resolutions comparable to EM. The single molecule force spectroscopy (SMFS) capacity of AFM also allowed us to perform a high-sensitivity mechanical profiling of the M. xanthus surface macromolecules. This analysis identified the surface cohesive substances as exopolysaccharides, providing the first nanomechanical evidence for their involvement in M. xanthus social behavior.