A novel immunity system for bacterial nucleic acid degrading toxins and its recruitment in various eukaryotic and DNA viral systems

Zhang, Dapeng; Iyer, Lakshminarayan M.; Aravind, L.

Identification of multiple distinct trafficking systems for toxins encoded in SUKH superfamily neighborhoods. We observed that in Gram-positive bacteria, proteins with the C-terminal nuclease typically possessed one of a set of several distinct domains at the extreme N-terminus of the protein ( Figure 5) . A significant subset of these could be unified using sequence profile searches with the PSI-BLAST and JACKHMMER programs to the WXG/ESAT6 superfamily of a-helical domains (65) . These domains are a specific signal recognized by the YueA-like ATPases of the HerA-FtsK superfamily that secrete them in an ATP-dependent manner (65, 66) . This indicated that the WXG/ESAT-6 domain-containing toxins in Gram-positive bacteria are extruded by YueA-like pumps using an ATP-dependent mechanism. A significant subset of toxin proteins from firmicutes possessed a distinctive N-terminal domain that could not be unified with any earlier known domain (a subset of these have been included in the erroneously annotated model Transposase_30 of PFAM; PF04740). Sequence searches showed that this domain possessed a conserved [LF]XG sequence motif and it was predicted to assume an a-helical bundle fold based on the multiple sequence alignment (Supplementary Data). We accordingly termed it the LXG domain ( Figure 5 ) and were able to unify it with the WXG domain by means of profile-profile comparisons with the HHpred program (P = 10 À7 ). Contextual analysis indicated that this domain is encoded by certain conserved gene-neighborhood across firmicutes, where it is associated with genes coding for a YueA-like HerA-FtsK superfamily protein pump and a small protein related to the S. aureus EsaC protein (gi: 282917938, Supplementary Data). Through profile-profile comparisons we showed that the EsaC-like superfamily is a bacterial version of the eukaryotic EVH1 peptide-binding domains with the PH-like fold (HHpred P-value: 10 À4 ) (67). These observations suggest that the LXG domain is comparable to the WXG/ESAT-6 domain, and is likely to utilize the ATP-dependent YueA pumps and the potential peptide-binding EsaC domain as partners for extrusion from the producing cell. The protein Srot_0310 (gi: 296392744) from the actinobacterium S. rotundus contains two copies of a distinct domain N-terminal to the GH-E nuclease domain ( Figure 5 ). This domain is also widely found in several actinobacteria at the N-termini of putative cell-surface proteins. Profileprofile comparisons suggested a possible relationship between these N-terminal domains and the WXG domain suggesting that it might be yet another representative of the WXG-like superfamily (P = 10 À4 ) and might utilize a similar ATP-dependent mechanism for its extrusion. A fourth group of proteins, restricted to certain firmicutes (e.g. S. aureus SACOL0281 protein; gi: 57652555), is typified by yet another N-terminal a-helical domain (LDXD in Figure 5 ) that is also found in domain architectural contexts very similar to the WXG and LXG domains. It is conceivable that this domain is comparable to them and functions similarly as a mediator of export via the HerA-FtsK superfamily pumps. Thus, a notable mode of export of nuclease toxins in Gram-positive bacteria appears to be via the ATP-dependent extrusion system, which while biochemically distinct from the TPS of the proteobacteria, is thematically comparable.