Woohoo, our paper on the discovery of 65 new phages dependent on conjugative plasmids is published in it's final form today in @NatureComms ! Here follows a thread (1/ ) link.springer.com/article/10…
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Natalia (@nquinoneso) already posted an excellent tweet thread summarising the findings of our earlier preprint (x.com/nquinoneso/status/1637…), so read that, and then let me elaborate on some of the exciting new data we added while this paper was in review.... (2/ )
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A major finding of our preprint was that though the phages we captured that were dependent on IncP conjugative plasmids were highly related genomically, they showed dramatic differences in their preference for different bacterial hosts (which all encode the same plasmid) (3/ )
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For example, the prototypical IncP plasmid-dependent phage PRD1 has a very generalist host range, and can infect most g-proteobacteria carrying an IncP plasmid. PRDcerulean is 95% identical to PRD1 but has a very narrow host range, mostly restricted to Pseudomonas (4/ )
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Naturally we wondered if defence systems native to these bacterial strains might explain any of the patterns. However, presence of sequence-specific defence systems (e.g. CRISPR & RM) did not account for the data, and we found no strong evidence defence systems are involved (5/ )
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After genetic detective work, we discovered that we could restore generalist host range to PRDcerulean by exchanging a single gene, encoding the holin protein (P35), with the PRD1 version! (6/ )
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Even further, we found single amino acid substitutions in the holin protein of PRDcerulean that could expand phage host range, and these mutations are associated with transmembrane domains (TMDs). Holin TMDs have been shown by Ry Young's lab to be critical to holin function (7/ )
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A second exciting development since the preprint was characterisation of the novel F plasmid-dependent phage, FtMidnight. This is the only tailed phage known to use the F pilus as a receptor (we don't know why but almost all conjugative pilus dep. phages are tail-less!) (9/ )
Apr 13, 2024 · 12:23 AM UTC
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Digging deeper into the FtMidnight genome sequence, we were able to annotate the putative distal tail region by structure guided homology search. Remarkably, we found that phages with related tail proteins to FtMidnight target the type 4 pilus (T4P)... (10/ )
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This is remarkable because despite the terribly-confusing nomenclature, the type 4 pilus (related to type 2 secretion systems) is a completely orthogonal contractile structure to the conjugative pilus (a type 4 secretion system)! (Fig adapted from doi.org/10.1038/emboj.2008.1…) (11/ )
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We're not yet sure which of FtMidnight's tail proteins interacts with the F pilus, but alignment of the putative distal tail proteins across the collection of related phage tails implicates gp18, as it is uniquely diverged away from the proteins of the T4P targeting phages (12/ )
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We are particularly excited about this finding, because it suggests conjugative plasmid (pilus) -dependency in phages can evolve from type 4 pilus targeting phages (which are quite common!)... (13/ )
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In fact, both other groups of F-plasmid dependent phages, the ssRNA Fiersviruses and the ssDNA Inoviruses have close relatives who use T4P receptors, suggesting evolutionary promiscuity for these two contractile structures in 3 unrelated groups of phages! 🤯(14/ )
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Stay tuned for more on these plasmid dependent phages in the future, along with other cool applications for the phage DisCo discovery platform coming soon 👀 (15/ )
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This work was a hugely enjoyable collaboration with @nquinoneso who just defended her PhD♥️, and we are grateful to our PI @baym for support and all co-authors for important contributions! Inc. @microMcCully @maxgmarin @EllieRand3 @RachelPetherbr1 & Jillian Paull! (16/ )
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Finally, I want to celebrate the four amazing undergraduate students from @bu_tweets and @SeeRCC : Alice Fan, Remi Martins Dosumu, Kay Paul and Cleo Sanchez Castaño who helped isolate some of these phages and coauthor this paper with us (17/ )
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