Signal transduction by two-component regulatory systems.<\/strong>\u00a0 The ability to respond to stimuli is often considered to be a key characteristic of life.\u00a0 Cells can detect new conditions, transduce that information into a usable form, and execute an appropriate response.\u00a0 Bacteria, archaea, fungi, and plants use two-component regulatory systems (TCSs) to regulate pathogenesis, antibiotic resistance, physiology, development, behavior, etc.\u00a0 Sensor kinases (the first component) detect stimuli and record this information in the form of phosphoryl groups.\u00a0 Response regulators (the second component) catalyze transfer of phosphoryl groups to themselves from sensor kinases (or from small molecules) to turn output function on, and from themselves to water to turn output function off.\u00a0 Inclusion of histidine-containing phosphotransfer (Hpt) domains enables more complex circuits with useful information processing and regulatory properties.\u00a0 Kinases and phosphatases accelerate response regulator autocatalytic reactions to achieve physiologically appropriate signaling speeds, but do not alter the intrinsic reaction mechanisms.\u00a0 The kinetics of biochemical signaling reactions are crucial to synchronize responses with stimuli, and can differ substantially for biological processes that operate on different timescales. \u00a0Our long-term goal is comprehensive understanding of signal transduction by TCSs, which we currently pursue by two general strategies:<\/p>\n An innovative research strategy for the genomics era.<\/strong>\u00a0 Genome sequencing presents both a challenge (a large gap between hundreds of thousands of known TCS proteins and what can be studied experimentally) and an opportunity (the availability of diverse and extensive sequence data).\u00a0 To elucidate general properties of TCS proteins, we investigate the consequences of sequence differences (rather than similarities) between the conserved domains that define sensor kinases or response regulators.\u00a0 For example, we have learned to alter the rate constants of response regulator phosphorylation and dephosphorylation reactions at least three orders of magnitude by changing specific variable residues in the active site, and understand the underlying molecular mechanisms.\u00a0 We use sequence data to drive experimental investigation of (i) self-catalyzed phosphorylation and dephosphorylation of response regulators, (ii) phosphotransfer reactions (including phosphorelays), (iii) phosphatase reactions, and (iv) synthetic biology applications.<\/p>\n Investigating chemotaxis phosphatases in\u00a0Vibrio cholerae<\/i><\/b>. \u00a0We are collaborating with Fitnat Yildiz (UC Santa Cruz) to investigate phosphatases involved in chemotaxis of\u00a0V. cholerae<\/i>, the causative agent of cholera. \u00a0The phosphorylation state of CheY-3 controls swimming behavior. \u00a0Purified CheZ has robust phosphatase activity toward phosphorylated CheY-3, whereas purified CheX exhibits modest activity under conditions tested so far. \u00a0In contrast, CheX has a larger effect than CheZ on swimming behavior of\u00a0V. cholerae<\/i>. \u00a0We have not yet identified the reasons for the discrepancy between our genetic and biochemical results. \u00a0We are also investigating the function and properties of a related DUF3334 protein, CheX2, using biochemical, bioinformatic, and genetic methods.<\/p>\n Park, J.H., Kennedy, E.N., Tripathi, S., Romp, A.B., Rubin, S.M., Bourret, R.B., & Yildiz, F. (2026) Distinct PlzC mechanisms integrate chemotaxis and c-di-GMP signaling to regulate Vibrio cholerae<\/i>\u00a0motility and biofilm formation. \u00a0Proc. Natl. Acad. Sci. U.S.A. <\/i>123<\/strong>, e2511740123<\/p>\n Bourret, R.B., Kennedy, E.N., Tamayo, R, & Foster, C.A. (2025) A clarifying perspective on bacterial pseudo-receiver domains.\u00a0\u00a0J. Bacteriol.<\/i>\u00a0207,<\/b>\u00a0e0026125. \u00a0Highlighted as an “Editor’s Pick”<\/p>\n Barr, S.A., Kennedy, E.N., McKay, L.S., Johnson, R.M., Ohr, R.J., Cotter, P.A., & Bourret, R.B. (2022) Phosphorylation chemistry of the Bordetella<\/em> PlrSR TCS and its contribution to bacterial persistence in the lower respiratory tract. Mol. Microbiol.<\/em> 119,<\/strong> 174-190.<\/p>\n Kennedy, E.N., Foster, C.A., Barr, S.A., & Bourret, R.B. (2022) General strategies for using amino acid sequence data to guide biochemical investigation of protein function. Biochem. Soc. Trans.<\/em> 50,<\/strong> 1847-1858.<\/p>\n Vass, L.R., Branscum, K.M., Bourret, R.B., & Foster, C.A. (2023) Analysis of CheW-like domains provides insights into organization of prokaryotic chemotaxis systems.\u00a0 Proteins<\/em>. 91<\/strong>, 315-329.<\/p>\n Vass, L.R., Branscum, K.M., Bourret, R.B., & Foster, C.A. (2022) Generalizable strategy to analyze domains in the context of parent protein architecture: A CheW case study. Proteins<\/em> 11<\/strong>, 1973-1986.<\/p>\n Kennedy, E.N., Barr, S.A., Liu, X., Vass, L.R., Liu, Y., Xie, Z., & Bourret, R.B. (2022) Azorhizobium caulinodans<\/em> chemotaxis is controlled by an unusual phosphorelay network.\u00a0 J. Bacteriol.<\/em> 204,<\/strong> e00527-21<\/p>\n Bourret, R.B., Foster, C.A., & Goldman, W.E. (2021) Predicted functional and structural diversity of receiver domains in fungal two-component regulatory systems. mSphere<\/em> 6,<\/strong> e00722-21.<\/p>\n Foster, C.A., Silversmith, R.E., Immormino, R.M., Vass, L.R., Kennedy, E.N., Pazy, Y., Collins, E.J., & Bourret, R.B. (2021) The role of position K+4 in phosphorylation and dephosphorylation reaction kinetics of the CheY response regulator. Biochemistry<\/em> 60,<\/strong> 2130-2151.<\/p>\n Bourret, R.B., Kennedy, E.N., Foster, C.A., Sepulveda, V.E., & Goldman, W.E. (2021) A radical reimagining of fungal two-component regulatory systems. Trends Microbiol.<\/em> 29,<\/strong> 883-893.<\/p>\n Department of Microbiology & Immunology<\/a><\/p>\n Lineberger Comprehensive Cancer Center (LCCC)<\/a><\/p>\n Program in Molecular and Cellular Biophysics<\/a><\/p>\n Biological and Biomedical Sciences Program (BBSP)<\/a><\/p>\nPublications<\/h3>\n
Affiliations<\/h2>\n