Navel type I restriction specificities through domain shuffling of HsdS subunits in Lactococcus lactis

University College Cork, Corcaigh, Munster, Ireland
Molecular Microbiology (Impact Factor: 4.42). 06/2000; 36(4):866-75. DOI: 10.1046/j.1365-2958.2000.01901.x
Source: PubMed


This study identifies a natural system in Lactococcus lactis, in which a restriction modification specificity subunit resident on a 6159 bp plasmid (pAH33) alters the specificity of a functional R/M mechanism encoded by a 20.3 kb plasmid, pAH82. The new specificity was identified after phenotypic and molecular analysis of a 26.5 kb co-integrate plasmid (pAH90), which was detected after bacteriophage challenge of the parent strain. Analysis of the regions involved in the co-integration revealed that two novel hybrid hsdS genes had been formed during the co-integration event. The HsdS chimeras had interchanged the C- and N-terminal variable domains of the parent subunits, generating two new restriction specificities. Comparison of the parent hsdS genes with other type I specificity determinants revealed that the region of the hsdS genes responsible for the co-integration event is highly conserved among lactococcal type I hsdS determinants. Thus, as hsdS determinants are widespread in the genus Lactococcus, new restriction specificities may evolve rapidly after homologous recombination between these genes. This study demonstrates that, similar to previous observations in Gram-negative bacteria, a Gram-positive bacterium can acquire novel restriction specificities naturally through domain shuffling of resident HsdS subunits.

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    • "LtrA is an essential splicing co-factor for Ll.LtrB that promotes the catalytically active tertiary conformation of the ribozyme [23]. Ll.LtrB is found on three L. lactis conjugative elements, the pAH90 plasmid [24] and two highly similar elements, the pRS01 plasmid [25] and the chromosomal sex factor (SF) [26]. In all cases, Ll.LtrB interrupts ltrB, a gene coding for relaxase (LtrB). "
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    ABSTRACT: Group II introns are RNA enzymes that splice themselves from pre-mRNA transcripts. Most bacterial group II introns harbour an open reading frame (ORF), coding for a protein with reverse transcriptase, maturase and occasionally DNA binding and endonuclease activities. Some ORF-containing group II introns were shown to be mobile retroelements that invade new DNA target sites. From an evolutionary perspective, group II introns are hypothesized to be the ancestors of the spliceosome-dependent nuclear introns and the small nuclear RNAs (snRNAs--U1, U2, U4, U5 and U6) that are important functional elements of the spliceosome machinery. The ability of some group II introns fragmented in two or three pieces to assemble and undergo splicing in trans supports the theory that spliceosomal snRNAs evolved from portions of group II introns. We used a transposon-based genetic screen to explore the ability of the Ll.LtrB group II intron from the Gram-positive bacterium Lactococcus lactis to be fragmented into three pieces in vivo. Trans-splicing tripartite variants of Ll.LtrB were selected using a highly efficient and sensitive trans-splicing/conjugation screen. We report that numerous fragmentation sites located throughout Ll.LtrB support tripartite trans-splicing, showing that this intron is remarkably tolerant to fragmentation. This work unveils the great versatility of group II intron fragments to assemble and accurately trans-splice their flanking exons in vivo. The selected introns represent the first evidence of functional tripartite group II introns in bacteria and provide experimental support for the proposed evolutionary relationship between group II introns and snRNAs.
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    • "Thus, the HsdSpIL2 is unlikely to form an active complex with the methylase subunit, despite having one conserved region. However, the hsdS gene of pIL2 might be the source of variation in specificity due to homologous recombination with other hsdS genes, resulting in formation of chimeric S-subunits [35]. In case of HsdSpIL6 and HsdSpIL7 proteins, both of them consist of conserved and variable regions typical for functional S-subunits. "
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    Full-text · Article · Jul 2011 · PLoS ONE
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    • "We do not know whether this integrase homolog is involved in recombination between the S loci as found in Mycoplasma pulmonis [39]. In Lactococcus lactis, two copies of the S genes on different plasmids, interact through homologous recombination and create two chimeric S genes for one RM system, resulting in shuffled recognition sequences [40]. "
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