Dissecting the bacterial type VI secretion system by a genome wide in silico analysis: what can be learned from available microbial genomic resources?

Abstract

Background: The availability of hundreds of bacterial genomes allowed a comparative genomic study of the Type VI Secretion System (T6SS), recently discovered as being involved in pathogenesis. By combining comparative and phylogenetic approaches using more than 500 prokaryotic genomes, we characterized the global T6SS genetic structure in terms of conservation, evolution and genomic organization.

Results: This genome wide analysis allowed the identification of a set of 13 proteins constituting the T6SS protein core and a set of conserved accessory proteins. 176 T6SS loci (encompassing 92 different bacteria) were identified and their comparison revealed that T6SS-encoded genes have a specific conserved genetic organization. Phylogenetic reconstruction based on the core genes showed that lateral transfer of the T6SS is probably its major way of dissemination among pathogenic and non-pathogenic bacteria. Furthermore, the sequence analysis of the VgrG proteins, proposed to be exported in a T6SS-dependent way, confirmed that some C-terminal regions possess domains showing similarities with adhesins or proteins with enzymatic functions.

Conclusion: The core of T6SS is composed of 13 proteins, conserved in both pathogenic and non-pathogenic bacteria. Subclasses of T6SS differ in regulatory and accessory protein content suggesting that T6SS has evolved to adapt to various microenvironments and specialized functions. Based on these results, new functional hypotheses concerning the assembly and function of T6SS proteins are proposed.

Figure 1

Genomic organization of the characterized T6SS loci. Genes are represented as arrows. Translated sequences of the most conserved proteins were aligned to the COG sequences and the hits are represented as coloured boxes. A unique colour is assigned to highly conserved COGs. Burkholderia mallei is closely related to B. pseudomallei, only this latter one is depicted.

Figure 2

T6SS genetic component frequencies. Frequencies were computed on the basis of all identified T6SS gene clusters except clusters encoded from closely related bacterial strains (in this case, only the clusters encoded in one strain has been taken into account).

Figure 3

T6SS consensual genomic organization. COGs are depicted as arrows. The colour scheme is as on Figure 1. When the order and the transcriptional orientation of two genes are conserved, the two corresponding COGs are linked. The more the genomic organization is conserved, the bolder the connectors. The percentages correspond to the link conservation frequencies. They were computed on the basis of all identified T6SS gene clusters except clusters encoded from closely related bacterial strains.

Figure 4

Relationship between phylogeny and T6SS gene content. Rows represent T6SS loci and columns represent protein functional classes (based on COG assignment). The tree on the left is the consensus phylogenetic tree (super-tree, see Materials and methods §5, manually rooted) whereas the upper dendrogram represents a hierarchical clustering of the phylogenetic profiles. Core T6SS conserved proteins are depicted on the right columns with the same color code as in Figures 1 and 3. Presence and absence of conserved accessory proteins (grey and light green) highlights the presence of sub-groups numbered from I to V. Letters in front of bacteria names correspond to the four groups proposed by Bingle and co-workers [14], '#' marks functional T6SS loci depicted in Table 1 and included in this figure. The two major sub-trees have been split and displayed separately for clarity. Light blue and light green highlighted groups are examples of close T6SS loci associated to bacteria with similar ecological niche. These two groups are associated to marine bacteria (sub-group V, blue) and plant associated bacteria (sub-group IV, green).

Figure 5

Relationship between phylogeny and T6SS gene content. Rows represent T6SS loci and columns represent protein functional classes (based on COG assignment). The tree on the left is the consensus phylogenetic tree (super-tree, see Materials and methods §5, manually rooted) whereas the upper dendrogram represents a hierarchical clustering of the phylogenetic profiles. Core T6SS conserved proteins are depicted on the right columns with the same color code as in Figures 1 and 3. Presence and absence of conserved accessory proteins (grey and light green) highlights the presence of sub-groups numbered from I to V. Letters in front of bacteria names correspond to the four groups proposed by Bingle and co-workers [14], '#' marks functional T6SS loci depicted in Table 1 and included in this figure. The two major sub-trees have been split and displayed separately for clarity. Light blue and light green highlighted groups are examples of close T6SS loci associated to bacteria with similar ecological niche. These two groups are associated to marine bacteria (sub-group V, blue) and plant associated bacteria (sub-group IV, green).