Discipline: Biological Sciences
Subcategory: Cell and Molecular Biology
Lam Vo - University of Tennessee at Knoxville
Co-Author(s): Gladys Alexandre, Ph.D at the University of Tennessee at Knoxville ; Tanmoy Mukherjee, Ph.D at the University of Tennessee at Knoxville
Bacterial chemotaxis is the biased movement of a motile bacterium in chemical gradients: toward regions with beneficial substances and away from regions with toxic substances. The best studied model for chemotaxis is Escherichia coli where a single chemotaxis, pathway that includes a single CheA histidine kinase and a single CheY response regulator coordinate changes in motility pattern during chemotaxis. However, unlike in E. coli, the genomes of most motile bacteria possess multiple CheAs and CheYs, but their exact role in chemotaxis is not well understood.
Azosprillum brasilense, the nitrogen-fixing bacterium that dwells in the soil, has been predicted to have 7 cheYs and 4 cheAs. This project aims to probe the function of these genes in modulating motility-dependent behaviors, including swimming, swarming, and biofilm formation. Mutant strains lacking genes encoding for these proteins were constructed using allelic exchange. These strains were characterized for swimming in liquid media as well as in media of increasing agar concentration to discriminate between polar flagellum dependent swimming (0.3 % agar) and lateral flagella dependent swarming (0.6% agar). In addition, protein-protein interactions between CheAs and CheYs were determined using bacterial-two hybrid assay (BACTH) and quantified using beta-galactosidase assay.
While each mutation caused defects in free-swimming in liquid media, there were significant differences in the ability of the mutant strains to swim in low agar medium and to swarm (student t-test, N=3, p < 0.05). All mutants that were unable to swim were also impaired in swarming. Together, these data suggest complex contribution of the multiple CheY homologs to polar and lateral flagella function with some CheYs controlling both types of motility apparatus. On the other hand, the mutants had distinct effects on biofilm formation ranging from no effect, inability to for biofilm to impaired and delayed biofilm. The range of chemotaxis and motility biases that the mutant exhibit together with their effect on biofilm formation suggest that change in motility during biofilm formation are orchestrated in a temporal manner and promoted by signaling through CheAs and CheYs, that still needs to be elucidated at greater resolution. These findings suggested a network of interaction of CheY homologs with each of the CheAs that provides a foundation for a working model for how these multiple systems control flagellar-function dependent behaviors. Future studies aim to decode specific interactions between multiple CheA, CheY, and both of flagella apparatuses using in vitro biochemical assay to form a model for these systems.
Wadhams H, George, Armitage P, Judith (2004). Making sense of it all: bacterial chemotaxis.
Nature Reviews Molecular Cell Biology 5, 1024-1037
Wuichet K & Zhulin IB (2010) Origins and diversification of a complex signal transduction system in prokaryotes. Sci Signal 3(128)
Funder Acknowledgement(s): I would like to thank Tanmoy Mukherjee for being my research mentor throughout this project. I would also like to thank Dr. Alexandre for being a very supportive principal investigator and advisor. This research was funded by NSF/REU grant to Gladys Alexandre, Ph.D.
Faculty Advisor: Dr. Gladys Alexandre, email@example.com
Role: When I first joined the lab, I was given this project by my faculty mentor Gladys Alexandre under the mentorship of Tanmoy Mukherjee. The project was already experimental designed by Tanmoy and my faculty mentor. I was the one performing the research, collecting data, and analyzing some part of the research (most of the result mentioned in this abstract is obtained by me). The motility pattern analysis of free-swimming bacteria in liquid media was an independent project performed by Tanmoy using homemade computerized tracking program; however, I collected data for this project also.