Peptidoglycan hydrolysis mediated by the amidase AmiC and its LytM activator NlpD is critical for cell separation and virulence in the phytopathogen Xanthomonas campestris

Abstract

SUMMARYThe essential stages of bacterial cell separation are described as the synthesis and hydrolysis of septal peptidoglycan (PG). The amidase, AmiC, which cleaves the peptide side-chains linked to the glycan strands, contributes critically to this process and has been studied extensively in model strains of Escherichia coli. However, insights into the contribution of this protein to other processes in the bacterial cell have been limited. Xanthomonas campestris pv. campestris (Xcc) is a phytopathogen that causes black rot disease in many economically important plants. We investigated how AmiC and LytM family regulators, NlpD and EnvC, contribute to virulence and cell separation in this organism. Biochemical analyses of purified AmiC demonstrated that it could hydrolyse PG and its activity could be potentiated by the presence of the regulator NlpD. We also established that deletion of the genes encoding amiC1 or nlpD led to a reduction in virulence as well as effects on colony-forming units and cell morphology. Moreover, further genetic and biochemical evidence showed that AmiC1 and NlpD affect the secretion of type III effector XC3176 and hypersensitive response (HR) induction in planta. These findings indicate that, in addition to their well-studied role(s) in cell separation, AmiC and NlpD make an important contribution to the type III secretion (T3S) and virulence regulation in this important plant pathogen.

Interplay between Peptidoglycan Biology and Virulence in Gram-Negative Pathogens

“…Meanwhile, in the genus Burkholderia, it has been shown that the inactivation of the periplasmic AmiC amidase causes the loss of daughter cell separation capacity, obviously entailing a severe impairment for motility, which was translated into the impossibility of causing infection in the rat model or colonizing the insect gut (68,69). Similar results for AmiC of the phytopathogen Xanthomonas campestris have been reported, together with negative effects on type III secretion system (T3SS) performance, obviously entailing an impairment of virulence (70). Nevertheless, at present it remains to be seen whether this impairment of the T3SS function could be related to defects in the insertion of its machinery through the PGN layer (caused by the AmiC absence), as will be displayed later for other proteins.…”

Repeated Phenotypic Evolution by Different Genetic Routes in Pseudomonas fluorescens SBW25

“…Accordingly, the next pair of mutations occur in an unrelated locus: nlpD ( pflu1301 ), which encodes a lipoprotein predicted to have a function in cell wall formation and cell separation in a range of bacteria (Stohl et al 2016; Lind et al 2017; Tsang et al. 2017; Yang et al. 2017).…”

The lipoprotein NlpD in Cronobacter sakazakii responds to acid stress and regulates macrophage resistance and virulence by maintaining membrane integrity

“…Previous studies have mostly considered that nlpD is related to envelop integrity and iron acquisition [ 44–46 ]. However, a few studies reported that nlpD affects virulence in the phytopathogens Xanthomonas [ 47 ] and Yersinia pestis [ 48 ]. Except in Yersinia pestis , however, the virulence mechanism of nlpD remains poorly understood.…”