Evolutionary thrift: mycobacteria repurpose plasmid diversity during adaptation of type VII secretion systems

Abstract

Mycobacteria have a distinct secretion system, termed type VII (T7SS), which is encoded by paralogous chromosomal loci (ESX) and associated with pathogenesis, conjugation, and metal homeostasis. Evolution of paralogous gene families is of interest because duplication is an important mechanism by which novel genes evolve, but there are potential conflicts between adaptive forces that stabilize duplications and those that enable evolution of new functions. Our objective was to delineate the adaptive forces underlying diversification of T7SS. Plasmid-borne ESX were described recently, and we found evidence that the initial duplication and divergence of ESX systems occurred on plasmids and was driven by selection for advantageous mutations. Plasmid conjugation has been linked to T7SS and type IV secretion systems (T4SS) in mycobacteria, and we discovered that T7SS and T4SS genes evolved in concert on the plasmids. We hypothesize that differentiation of plasmid ESX helps to prevent conjugation among cells harboring incompatible plasmids. Plasmid ESX appear to have been repurposed following migration to the chromosome, and there is evidence of positive selection driving further differentiation of chromosomal ESX. We hypothesize that ESX loci were initially stabilized on the chromosome by mediating their own transfer. These results emphasize the diverse adaptive paths underlying evolution of novelty, which in this case involved plasmid duplications, selection for advantageous mutations in the mobile and core genomes, migration of the loci between plasmids and chromosomes, and lateral transfer among chromosomes. We discuss further implications for the choice of model organism to study ESX functions in Mycobacterium tuberculosis.

Comprehensive in silico survey of the Mycolicibacterium mobilome reveals an as yet underexplored diversity

“…None of the putative plasmid sequences branched into chromosomal ESX clades, grouping mainly with sequences of known plasmids from different species of Mycobacteriaceae, which gives confidence that these sequences belong to mobile elements and reinforces the hypothesis of T7SS mobility [13]. Regarding the ESX types most commonly identified here, ESX-2 and ESX-5, they appear to be exclusive to plasmids, as there are no reports of them on Mycolicibacterium chromosomes [11][12][13]. While Mycolicibacterium plasmids carrying ESX-2 are common, those with ESX-5 have rarely been observed [13].…”

“…In addition, an association of T4SS-like genes with the T7SS has been observed, mainly in plasmids harbouring ESX-2 and ESX-5, as most of them carried vir B4-like, vir D4-like and tcp C-like genes close to the T7SS loci, resembling a conjugation-related locus [13]. None of the putative plasmid sequences branched into chromosomal ESX clades, grouping mainly with sequences of known plasmids from different species of Mycobacteriaceae , which gives confidence that these sequences belong to mobile elements and reinforces the hypothesis of T7SS mobility [13]. Regarding the ESX types most commonly identified here, ESX-2 and ESX-5, they appear to be exclusive to plasmids, as there are no reports of them on Mycolicibacterium chromosomes [11–13].…”

“…(iii) Circularity inference of the Mycolicibacterium contigs: assembly programs only produce linear contigs, so if a single contig has sufficient coverage with reads overlapping its ends, it is evidence of its circularity [ 35 ]. (iv) Searches for T7SSs related to plasmids because plasmid T7SS sequences differ from those encoded on chromosomes [ 13 ]. The T7SS core proteins (EccA, EccB, EccC, EccD, EccE and MycP) were surveyed by the hmmsearch program [ 34 ] using the hmm profiles listed in Table S4.…”

“…We also observed in 40/156 (~25 %) of the predicted plasmids clusters of genes that resemble the T7SS. The T7SS is a common feature in Mycobacteriaceae plasmids, and in fact, they played a major role in the evolution and radiation of this secretion system in this bacterial family [11][12][13]. Among the predicted plasmids, those harbouring the T7SS had a median size (~117 kb) larger than plasmids that did not encode the T7SS (~21 kb) (P <0.05), and all ESX types were represented, except ESX-4, with most sequences encoding ESX-2 and ESX-5 (Fig.…”

“…Despite this, plasmids have been associated with the evolution and dissemination of the major specialized secretion system among hundreds of mycobacteria species, namely the type-VII secretion system (T7SS or ESX). This secretion system is encoded by six paralogous loci (ESX-1, -2, -3, -4, -4- bis and -5) in plasmids and chromosomes [11–13]. To date, most of the plasmids identified in this family ( n =94) belong to the genus Mycobacterium (https://www.ncbi.nlm.nih.gov/genome/browse#!/plasmids/mycobacterium), probably due to its clinical relevance.…”