Involvement of the LlaKR2I methylase in expression of the AbiR bacteriophage defense system in Lactococcus lactis subsp lactis biovar diacetylactis KR2

The native lactococcal plasmid, pKR223, from Lactococcus lactis subsp. lactis biovar diacetylactis KR2 encodes two distinct bacteriophage-resistant mechanisms, the L1aKR21 restriction and modification (R/M) system and the abortive infection (Abi) mechanism, AbiR, that impedes bacteriophage DNA replication. This study completed the characterization of AbiR, revealing that it is the first Abi system to be encoded by three genes, abiRa, abiRb, and abiRc, arranged in an operon and that it requires the methylase gene from the L1aKR21 R/M system. An analysis of deletion and insertion clones demonstrated that the AbiR operon was toxic in L. lactis without the presence of the L1aKR21 methylase, which is required to protect L. lactis from AbiR toxicity. The novelty of the AbiR system resides in its original gene organization and the unusual protective role of the L1aKR21 methylase. Interestingly, the AbiR genetic determinants are flanked by two IS982 elements generating a likely transposable AbiR composite. This observation not only substantiated the novel function of the L1aKR21 methylase in the AbiR system but also illustrated the evolution of the L1aKR21 methylase toward a new and separate cellular function. This unique structure of both the L1aKR21 R/M system and the AbiR system may have contributed to the evolution of the L1aKR21 methylase toward a novel role comparable to that of the cell cycle-regulated methylases that include Dam and CcrM methylases. This new role for the L1aKR21 methylase offers a unique snapshot into the evolution of the cell cycle-regulated methylases from an existing R/M system.