The Caulobacter NtrB-NtrC two-component system bridges nitrogen assimilation and cell development [RNA-seq]
Caulobacter species are dimorphic Gram-negative bacteria that inhabit diverse terrestrial and aquatic ecosystems. A suite of molecular sensory systems enables these microbes to control growth, development, and reproduction in response to levels of essential elements, including carbon, nitrogen, and phosphorus. Although the enhancer binding protein NtrC and its cognate sensor histidine kinase NtrB are well-established regulators of bacterial nitrogen assimilation, their precise functions in Caulobacter metabolism and cell development remained largely undefined. Deletion of C. crescentus ntrC slowed cell growth in complex medium, while ntrB and ntrC were essential for growth when ammonium was the sole nitrogen source due to their requirement for glutamine synthase (glnA) expression. Random transposition of a conserved IS3-family mobile genetic element frequently rescued the growth defect of ntrC mutant strains by restoring glnBA transcription, revealing a possible role for IS3 transposition in shaping the evolution of Caulobacter populations during nitrogen limitation. We further defined the NtrC regulon through transcriptomic and ChIP-seq analyses, which demonstrated the role of NtrC as both a transcriptional activator and repressor. The chromosome of C. crescentus harbors dozens of direct binding sites for NtrC, with a significant proportion located near genes involved in polysaccharide biosynthesis. Numerous NtrC binding sites align with those of GapR, an essential protein involved in chromosome organization, and MucR1, a cell cycle regulator. This observation suggests that NtrC participates in the regulation of cell cycle and cell development. Indeed, loss of NtrC function led to elongated polar stalks and elevated synthesis of cell envelope polysaccharides. These developmental defects were restored by supplementing media with glutamine or by ectopic expression of the glnBA operon. This study establishes a regulatory connection between NtrC, nitrogen metabolism, polar morphogenesis, and envelope polysaccharide synthesis in Caulobacter. Overall design: To investigate the NtrC regulon in Caulobacter by creating an in-frame knockout of ntrC To investigate effects of glutamine supplementation on global expression in both WT and ?ntrC  by growing  these Caulobacter strains in peptone yeast extract (PYE) complex medium supplemented with 9.3 mM (final concentration) glutamine We performed gene expression analysis from RNA-seq data of WT and ?ntrC Caulobacter strains were grown in PYE or PYE supplemented with 9.3 mM (final concentration) glutamine to log phase Gene expression profiles across strains (i.e., WT, ?ntrC) and/or across medium condidtions (i.e., PYE, PYE supplemented with 9.3 mM glutamine) were compared