Title: Regulatory mechanisms controlling temperature-dependent transcription of flagellar motility genes in Listeria monocytogenes
Description: Flagellar motility is the primary mechanism of locomotion used by most bacterial species. Although flagella confer an advantage to many bacterial pathogens for colonization during infection, bacterial flagellins also stimulate host innate immune responses. Listeria monocytogenes is a facultative intracellular pathogen that represses transcription of flagellar motility genes at physiological temperatures (37°C and above). This dissertation describes experiments that further characterize the regulatory mechanisms required for temperature-dependent transcription of flagellar motility genes in L. monocytogenes and identifies the temperature-dependent mechanism that governs temperature specificity of flagellar motility gene transcription in L. monocytogenes . The MogR repressor and the DegU response regulator had been previously identified as transcriptional regulators of flagellar motility genes in L. monocytogenes . MogR represses transcription of all flagellar motility genes in a non-hierarchal manner by binding directly to flagellar promoter region DNA. In this dissertation, we identified a DegU-regulated bifunctional protein (GmaR) that functions both as a MogR anti-repressor and a flagellin glycosyltransferase. GmaR is expressed only at temperatures below 37°C. As an anti-repressor, GmaR forms a stable protein-protein complex with MogR, inhibiting MogR DNA binding activity at low temperatures and permitting flagellar motility gene transcription. We determined that DegU constitutively activates transcription of gmaR , in a phosphorylation-independent and temperature-independent manner.

We also demonstrated that GmaR has O-linked N-acetylglucosamine transferase (OGT) activity for flagellin, which established GmaR as the first flagellin glycosyltransferase to be identified and characterized in a Gram-positive organism, and the first known example of a bifunctional protein that transcriptionally regulates expression of its enzymatic substrate. Finally, in addition to being an anti-repressor and a glycosyltransferase, GmaR is also a protein thermometer. GmaR undergoes a temperature-dependent conformational change at 37°C, that affects the formation of the MogR:GmaR anti-repression complex and alters GmaR stability. GmaR is the first known example of a protein thermometer that functions as an anti-repressor. This unique thermal-sensing mechanism transduces temperature signals into a transcriptional response that controls temperature-dependent expression of flagella in L. monocytogenes.