The Stealth Episome: Suppression of Gene Expression on the Excised Genomic Island PPHGI-1 from Pseudomonas syringae pv. phaseolicola

Abstract

Pseudomonas syringae pv. phaseolicola is the causative agent of halo blight in the common bean, Phaseolus vulgaris. P. syringae pv. phaseolicola race 4 strain 1302A contains the avirulence gene avrPphB (syn. hopAR1), which resides on PPHGI-1, a 106 kb genomic island. Loss of PPHGI-1 from P. syringae pv. phaseolicola 1302A following exposure to the hypersensitive resistance response (HR) leads to the evolution of strains with altered virulence. Here we have used fluorescent protein reporter systems to gain insight into the mobility of PPHGI-1. Confocal imaging of dual-labelled P. syringae pv. phaseolicola 1302A strain, F532 (dsRFP in chromosome and eGFP in PPHGI-1), revealed loss of PPHGI-1::eGFP encoded fluorescence during plant infection and when grown in vitro on extracted leaf apoplastic fluids. Fluorescence-activated cell sorting (FACS) of fluorescent and non-fluorescent PPHGI-1::eGFP F532 populations showed that cells lost fluorescence not only when the GI was deleted, but also when it had excised and was present as a circular episome. In addition to reduced expression of eGFP, quantitative PCR on sub-populations separated by FACS showed that transcription of other genes on PPHGI-1 (avrPphB and xerC) was also greatly reduced in F532 cells harbouring the excised PPHGI-1::eGFP episome. Our results show how virulence determinants located on mobile pathogenicity islands may be hidden from detection by host surveillance systems through the suppression of gene expression in the episomal state.

Inactivation of PbTopo IIIβ causes hyper‐excision of the Pathogenicity Island HAI2 resulting in reduced virulence of Pectobacterium atrosepticum

“…In planta excision of PPHGI‐1, a HAI in P. syringae pv. phaseolicola , results in lower transcription of several genes on the island including the key avirulence gene hopAR1 (Godfrey et al ., 2011). Furthermore, the presence of the conjugative transposon CTnDOT in the chromosome of Bacteroides thetaiotaomicron alters chromosomal gene expression in its bacterial host (Moon et al ., 2007).…”

“…Certainly, induced excision of PPHGI‐1 in P. syringae pv. phaseolicola in planta results in the suppression of gene expression on the island, almost silencing expression of the key avirulence gene h opAR1 (Godfrey et al ., 2011). Inactivation of Pb Topo IIIβ results in induced transcription of several genes present on HAI2 both in vitro and in planta.…”

Genetics and Functional Genomics of the Pseudomonas fluorescens Group

“…Whether some or all of them are capable of excision from their genomic location is unclear. Functional studies similar to those done for islands in P. aeruginosa (Mathee et al 2008) and P. syringae (Godfrey et al 2013) are required if we are to fully understand the data generated by DNA sequencing.…”

The influence of the accessory genome on bacterial pathogen evolution

“…87 Moreover, it raises the question of whether other recombinases (e.g., those of ICEs, GIs, IS elements and transposons) are also affected by conjugation and regulated by LexA and the SOS response-certainly, stress affects P. syringae PPHGI-1 integrase expression. [58][59][60]88,89 Integrons have been found in a wide range of non-pathogenic and pathogenic bacteria in environments ranging from marine to terrestrial organisms and niches. 73,77,83,[90][91][92][93][94][95][96][97][98][99][100][101] Two types of integrons, the chromosomal integron (CI) and mobile integron (MI) have been identified: 70 CIs appear to represent a "core" gene capture system in Gram negative bacteria to enable genome flexibility for adaptation to diverse environments.…”