Here we report the discovery of two additional gene clusters that together with the ids operon comprise a network of self-recognition genes (). One locus, idr, encodes proteins necessary for merging with the parent BB2000 strain, while the other locus, tss, encodes a type VI secretion system that mediates export of Ids and Idr proteins. Significantly, we found that multiple Ids and Idr proteins are exported from the cell, including the self-identity determinant protein, IdsD.
FIG 4 Model for Ids and Idr functional roles in self-recognition. (A) Functional flow chart for the roles of the Ids, Idr, and T6S proteins in self-recognition and territorial behaviors. A subset of Ids and Idr proteins are primarily exported via a shared T6S (more ...)
The Idr and Ids proteins represent two separate mechanisms for self-recognition. Export of the Idr proteins is independent of the ids
gene cluster, and likewise, export of the Ids proteins is not dependent on the idr
gene cluster. Moreover, strains with mutations in the ids
genes have different phenotypes in intraspecies competitions. The Ids proteins, which are needed only for competition with the parent strain, encode strain-specific self-identity determinants (15
). Interestingly, the idrD
gene contains rhs
sequences, which are commonly found in bacterial species. Recent research has implicated that these rhs
-encoded proteins, as well as proteins involved in contact-dependent inhibition, such as CdiA in E. coli
, encode toxin elements in the C-terminal domain (6
). However, the rhs
-containing sequences may also encode adhesion molecules, because they share some sequence similarity to YD-repeat-containing teneurin proteins (11
). Either of these proposed idrD
functions could explain why the idr
genes are required for increased competition (and/or population migration) against foreign strains.
Indeed, we demonstrate that the self-recognition capability of P. mirabilis
provides a competitive advantage for the population specifically on surfaces. We did not observe similar advantages when the wild type was grown with self-recognition mutant strains in liquid (see the supplemental material). Growth on surfaces induces a broad developmental change in P. mirabilis
where increased cell-cell contact yields increased population-wide coordination that is integral to migration and outward expansion of the swarm (reviewed in references 20
). Perhaps the behavior of self-recognition is most beneficial in environments where social interactions are more frequent and thus potentially more impactful.
Our research supports a model in which P. mirabilis
self-recognition involves the display of self-identity proteins that are likely interpreted via a direct physical interaction with other cells; this communication then yields a self-versus-nonself assessment that guides whether boundaries are formed between populations (). Some self-identity proteins are likely displayed on or near the cell surface, as physical contact between cells is required for boundary formation (22
). This extracellular exposure may serve to communicate a cell’s identity represented by the self-identity determinant molecules IdsD and IdsE during interactions with neighboring cells (15
). Indeed, Ids and Idr proteins are transported out of the cell via the T6S system and either are transported into the neighboring cell or localize on the cell surface (see the supplemental material). However, we have not yet found evidence for the transfer of self-recognition proteins into a neighboring cell.
We propose that boundary formation can result from the actions of a single population, which has queried on a cell-cell level whether the neighboring cell is self. For each population of P. mirabilis strain BB2000, “self” is defined by the combined actions of the Ids and Idr pathways. Self-recognition occurs when both the expected cognate Ids and Idr proteins are present in (or on) the neighboring cell, ultimately resulting in merging with the neighboring swarm (). In contrast, we predict that boundary formation results from the absence of the cognate Ids and Idr self determinants in the neighboring cell (). Both the Ids and Idr proteins likely initiate downstream signaling pathways that are altered depending on whether the interactions are with cognate or noncognate Ids and Idr proteins, respectively.
This two-part network for self-recognition appears analogous to aspects of the innate immune system and indeed has many parallels to the immune surveillance of natural killer (NK) cells. In current models for NK cell activity, the presence of self cells (i.e., of one’s own organism) is conveyed by the combined detection of two surface receptors (an activation receptor ligand and class I major histocompatibility complex [MHC]), resulting in no killing of the self cell. In contrast, the absence of either receptor leads to the NK cell’s determination of an absence of self and the subsequent killing of the nonself (or receptor-deficient) cell, as reviewed in reference 23
. Intriguingly, these results in P. mirabilis
further support the idea that cellular self-recognition is a behavior shared among organisms at many levels of biological complexity.
While the capability for self-recognition is broadly found, it remains unclear why and how bacteria utilize this ability. In P. mirabilis
, self-recognition is necessary for territorial expansion when one population interacts with competing nonisogenic populations. Recently, other research groups have shown that type VI secretion systems confer a fitness advantage in interbacterial and interkingdom competitions, likely through transport of small peptides, but their role in intraspecies interactions is only beginning to emerge (3
). Our demonstration that a T6S system functions directly in self-recognition-dependent territoriality expands the set of known applications for this widely conserved export machinery. We still need to explore the mechanisms of T6S in P. mirabilis
and its relative functional capabilities compared to T6S systems described in other bacteria. Importantly, we still need to understand the dynamics of Idr and Ids protein-protein interactions within and between cells. Indeed, the Ids, Idr, and T6S proteins together form a mechanistic foundation for examining the basic biological phenomena of territoriality and self-recognition in a bacterial model system.