Abstract
Outer membrane vesicles (OMVs), released from Gram-negative bacteria, have been attributed to intra- and interspecies communication and pathogenicity in diverse bacteria. OMVs carry various components including genetic material, toxins, signaling molecules, or proteins. Although the molecular mechanism(s) of cargo delivery is not fully understood, recent studies showed that transfer of the OMV content to surrounding cells is mediated by selective interactions. Here, we show that the phytopathogen Agrobacterium tumefaciens, the causative agent of crown gall disease, releases OMVs, which attach to the cell surface of various Gram-negative bacteria. The OMVs contain the conserved small lipoprotein Atu8019. An atu8019-deletion mutant produced wildtype-like amounts of OMVs with a subtle but reproducible reduction in cell-attachment. Otherwise, loss of atu8019 did not alter growth, susceptibility against cations or antibiotics, attachment to plant cells, virulence, motility, or biofilm formation. In contrast, overproduction of Atu8019 in A. tumefaciens triggered cell aggregation and biofilm formation. Localization studies revealed that Atu8019 is surface exposed in Agrobacterium cells and in OMVs supporting a role in cell adhesion. Purified Atu8019 protein reconstituted into liposomes interacted with model membranes and with the surface of several Gram-negative bacteria. Collectively, our data suggest that the small lipoprotein Atu8019 is involved in OMV docking to specific bacteria.
Highlights
The release of membrane vesicles (MVs) is ubiquitous among bacteria
To study outer membrane vesicles (OMVs) release and composition from A. tumefaciens, we isolated OMVs from cell-free supernatants of early stationary phase cultures grown in LB medium
SDS-PAGE analysis of the OMV preparation revealed a strong reduction of the number of protein bands in comparison with whole-cell lysates (CP) and an enrichment of proteins of about 13 to 35 kDa (Figure 1B)
Summary
MVs from Gram-negative bacteria are usually termed outer membrane vesicles (OMVs) They can be formed by budding off the outer membrane (OM) after cargo enclosure by various mechanisms (Hoekstra et al, 1976; Zhou et al, 1998; Lee et al, 2008; Bonnington and Kuehn, 2014; Kim et al, 2015; Schwechheimer and Kuehn, 2015; Toyofuku et al, 2017; Toyofuku, 2019). Recent studies provided evidence that MVs selectively interact with bacteria in order to transfer their content to the target cells (MacDonald and Beveridge, 2002; Lin et al, 2017; Tashiro et al, 2017; Brameyer et al, 2018). Some LPPs are surfaceexposed even in heterologous host cells, suggesting an additional, and yet unidentified universally conserved mechanism (Arnold et al, 2014; Zückert, 2014; Konovalova and Silhavy, 2015; Hooda et al, 2016; Wilson and Bernstein, 2016; Fantappie et al, 2017)
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