Flagellar motility is essential for many pathogenic bacteria to promote infection of hosts to result in disease. In addition, flagellar motility allows many environmental bacteria to reach specific niches and nutrients for optimal growth. The structure and function of a prototypic flagellar nanomachine is largely derived from cumulative studies of peritrichous flagella from Salmonella species and Escherichia coli. A typical flagellum is composed of a flagellar motor that extends from the inner and outer membranes, a surface-localized hook, and an extracellular filament that rotates. The flagellar motor is composed of stator complexes in the inner membrane that associate with rotor and switch components at the base of the flagellum and a rod that penetrates the peptidoglycan and the outer membrane. The stators generate an ion gradient to power the rotor to turn the rod and the filament, which results in swimming motility. The flagellar motors of polarly-flagellated bacteria contain these core components, but also possess additional structural components that are essential for motor function. By using electron cryo-tomography, we have identified three disk structures associated with the polar flagellar motor of Campylobacter jejuni, a leading cause of gastroenteritis in humans in the United States and in other countries throughout the world, which are essential for promoting motility. We have identified two proteins, FlgP and Cjj0413, which compose two of these motor disk structures. Deletion of either protein abolishes motor function and motility in C. jejuni. Because homologues of these proteins are found in other polar flagellates that also produce disk-like structures associated with their respective flagellar motors, we believe that we have identified components universally required for function of polar flagellar motors. The objectives of this proposal are to use C. jejuni as a model system to understand construction and function of flagellar motors as they relate to medically-important polar-flagellated bacterial pathogens such as Vibrio, Pseudomonas, and Helicobacter species.
In Aim 1, we will perform experiments to identify other proteins of C. jejuni forming motor disk structures and proteins specifically required for flagellar motor function.
In Aim 2, we will analyze essential residues of FlgP required for formation of the basal motor disk and nucleation of the other motor disks. In addition, we will analyze cytoplasmic and periplasmic domains of Cjj0413 and their requirements for polymerization of the proximal disk and motility. Accomplishment of these aims will aid in understanding: 1) unique components required for a functional polar flagellar motor;2) alternative mechanisms for the construction and function of polar flagellar motors that differ from peritrichous motors;and 3) diversity amongst motile bacteria for how flagellar nanomachines function to promote swimming motility across bacterial species, which is essential for optimal growth of many bacteria in hosts and in environments.
Flagellar motility in many bacteria is powered by a flagellar motor that facilitates rotational movement of an extracellular filament. Recent structural investigations have revealed that flagellar motors of polarly-flagellated bacteria are more complex than those of peritrichous bacteria and contain additional structural components essential for promoting motility. The proposed research will use Campylobacter jejuni as a model system to identify and characterize components forming three novel disk structures associated with a polar flagellar motor that are essential for motility, which will provide insight into understanding how flagellar motors function in a broad range of polar-flagellated bacteria.
|Beeby, Morgan; Ribardo, Deborah A; Brennan, Caitlin A et al. (2016) Diverse high-torque bacterial flagellar motors assemble wider stator rings using a conserved protein scaffold. Proc Natl Acad Sci U S A 113:E1917-26|
|Gulbronson, Connor J; Ribardo, Deborah A; Balaban, Murat et al. (2016) FlhG employs diverse intrinsic domains and influences FlhF GTPase activity to numerically regulate polar flagellar biogenesis in Campylobacter jejuni. Mol Microbiol 99:291-306|
|Luethy, Paul M; Huynh, Steven; Parker, Craig T et al. (2015) Analysis of the activity and regulon of the two-component regulatory system composed by Cjj81176_1484 and Cjj81176_1483 of Campylobacter jejuni. J Bacteriol 197:1592-605|
|Barrero-Tobon, Angelica M; Hendrixson, David R (2014) Flagellar biosynthesis exerts temporal regulation of secretion of specific Campylobacter jejuni colonization and virulence determinants. Mol Microbiol 93:957-74|
|Kendall, John J; Barrero-Tobon, Angelica M; Hendrixson, David R et al. (2014) Hemerythrins in the microaerophilic bacterium Campylobacter jejuni help protect key iron-sulphur cluster enzymes from oxidative damage. Environ Microbiol 16:1105-21|
|Boll, Joseph M; Hendrixson, David R (2013) A regulatory checkpoint during flagellar biogenesis in Campylobacter jejuni initiates signal transduction to activate transcription of flagellar genes. MBio 4:e00432-13|
|Kazmierczak, Barbara I; Hendrixson, David R (2013) Spatial and numerical regulation of flagellar biosynthesis in polarly flagellated bacteria. Mol Microbiol 88:655-63|
|Beeby, Morgan; Gumbart, James C; Roux, BenoÃ®t et al. (2013) Architecture and assembly of the Gram-positive cell wall. Mol Microbiol 88:664-72|