Genome sequence data has determined the duodenal
ulcer promoting (dupAgene to be encoded within a cluster of vir homologous
type IV secretion system T4SS) genes in the plasticity zones of several H.
pylori strains. Sequence identity and the presence of characteristic sequence
motifs indicates that DupA may be the VirB4 ATPase component of the T4SS,
however the function of the T4SS, recently termed Tfs4, and the identity of
its secretion substrates are unknown.
In this study, we aimed to assess the
protein-protein interactions mediated by a tfs4-encoded VirD2-like protein
using the yeast two-hybrid (Y2H) system. VirD2 proteins are relaxases
typically involved in conjugation or interkingdom DNA transfer in association
with several other proteins collectively referred to as the relaxosome. A homologue
of one other relaxosome protein, VirC1 is also present in tfs4.
In a candidate approach, a pairwise Y2H
interaction screen determined that VirD2 interacted with itself, VirC1 and an
unknown protein encoded adjacent to VirC1.
Interactions were generally weak indicating a
likely requirement for stabilising factors inherent to a relaxosome complex.
An interaction was not observed with a VirD4 coupling protein. In a secondary
approach, a high titre genomic library has been constructed from a tfs3 +
/tfs4 + /cag+ H. pylori clinical strain and is in the process of being
screened to deﬁne the entire repertoire of T4SS proteins that
interact with VirD2 and VirD4.
Our preliminary observations are consistent with
known interactions in other T4SSs and suggest that Tfs4 may function in DNA
transfer to a host cell.