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Salmonella Genomic Island 1 requires a self-encoded small RNA for mobilization.

ABSTRACT: The SGI1-family elements that are specifically mobilized by the IncA- and IncC-family plasmids are important vehicles of antibiotic resistance among enteric bacteria. Although SGI1 exploits many plasmid-derived conjugation and regulatory functions, the basic mobilization module of the island is unrelated to that of IncC plasmids. This module contains the oriT and encodes the mobilization proteins MpsA and MpsB, which belong to the tyrosine recombinases and not to relaxases. Here we report an additional, essential transfer factor of SGI1. This is a small RNA deriving from the 3'-end of a primary RNA that can also serve as mRNA of ORF S022. The functional domain of this sRNA named sgm-sRNA is encoded between the mpsA gene and the oriT of SGI1. Terminator-like sequence near the promoter of the primary transcript possibly has a regulatory function in controlling the amount of full-length primary RNA, which is converted to the active sgm-sRNA through consecutive maturation steps influenced by the 5'-end of the primary RNA. The mobilization module of SGI1 seems unique due to its atypical relaxase and the newly identified sgm-sRNA, which is required for the horizontal transfer of the island but appears to act differently from classical regulatory sRNAs.

SUBMITTER: Nagy I

PROVIDER: S-EPMC9299015 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:IncC conjugative plasmids and Salmonella genomic island 1 (SGI1) and relatives are frequently associated with multidrug resistance of clinical isolates of pathogenic Enterobacteriaceae. SGI1 is specifically mobilized in trans by IncA and IncC plasmids (commonly referred to as A/C plasmids) following its excision from the chromosome, an event triggered by the transcriptional activator complex AcaCD encoded by these helper plasmids. Although SGI1 is not self-transmissible, it carries three genes, traNS, traHS and traGS, coding for distant homologs of the predicted mating pore subunits TraNC, TraHC and TraGC, respectively, encoded by A/C plasmids. Here we investigated the regulation of traNS and traHGS and the role of these three genes in the transmissibility of SGI1. Transcriptional fusion of the promoter sequences of traNS and traHGS to the reporter gene lacZ confirmed that expression of these genes is inducible by AcaCD. Mating experiments using combinations of deletion mutants of SGI1 and the helper IncC plasmid pVCR94 revealed complex interactions between these two mobile genetic elements. Whereas traNC and traHGC are essential for IncC plasmid transfer, SGI1 could rescue null mutants of each individual gene revealing that TraNS, TraHS and TraGS are functional proteins. Complementation assays of individual traC and traS mutants showed that not only do TraNS/HS/GS replace TraNC/HC/GC in the mating pore encoded by IncC plasmids but also that traGS and traHS are both required for SGI1 optimal transfer. In fact, remodeling of the IncC-encoded mating pore by SGI1 was found to be essential to enhance transfer rate of SGI1 over the helper plasmid. Furthermore, traGS was found to be crucial to allow DNA transfer between cells bearing IncC helper plasmids, thereby suggesting that by remodeling the mating pore SGI1 disables an IncC-encoded entry exclusion mechanism. Hence traS genes facilitate the invasion by SGI1 of cell populations bearing IncC plasmids.

Project description:BACKGROUND: The Salmonella genomic island 1 (SGI1) is a 42.4 kb integrative mobilizable element containing several antibiotic resistance determinants embedded in a complex integron segment In104. The numerous SGI1 variants identified so far, differ mainly in this segment and the explanations of their emergence were mostly based on comparative structure analyses. Here we provide experimental studies on the stability, entrapment and variant formation of this peculiar gene cluster originally found in S. Typhimurium. METHODOLOGY/PRINCIPAL FINDINGS: Segregation and conjugation tests and various molecular techniques were used to detect the emerging SGI1 variants in Salmonella populations of 17 Salmonella enterica serovar Typhimurium DT104 isolates from Hungary. The SGI1s in these isolates proved to be fully competent in excision, conjugal transfer by the IncA/C helper plasmid R55, and integration into the E. coli chromosome. A trap vector has been constructed and successfully applied to capture the island on a plasmid. Monitoring of segregation of SGI1 indicated high stability of the island. SGI1-free segregants did not accumulate during long-term propagation, but several SGI1 variants could be obtained. Most of them appeared to be identical to SGI1-B and SGI1-C, but two new variants caused by deletions via a short-homology-dependent recombination process have also been detected. We have also noticed that the presence of the conjugation helper plasmid increased the formation of these deletion variants considerably. CONCLUSIONS/SIGNIFICANCE: Despite that excision of SGI1 from the chromosome was proven in SGI1(+)Salmonella populations, its complete loss could not be observed. On the other hand, we demonstrated that several variants, among them two newly identified ones, arose with detectable frequencies in these populations in a short timescale and their formation was promoted by the helper plasmid. This reflects that IncA/C helper plasmids are not only involved in the horizontal spreading of SGI1, but may also contribute to its evolution.

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Salmonella Genomic Island 1 requires a self-encoded small RNA for mobilization.

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