Novel Mechanisms Regulating Expression of Streptococcus Mutans Genomic Island TnSmu2

Objectives: Despite being present in only a subset of isolates, bacterial genomic islands (GI) are often significant contributors to pathogenicity. The GI designated TnSmu2 found in Streptococcus mutans UA159 contains the mub operon (SMU.1334c to SMU.1349) encoding genes involved in production of a nonribosomal peptide and polyketide. As part of the core genome, SMU.1655c encodes a peptide and is situated in an operon for serine metabolism (ser). This study assesses the contribution of SMU.1655c to the pathogenesis of the caries pathogen.
Methods: RNA-Seq and RT-PCR were performed to assess gene expression, and various mutants were characterized in growth rate, competence and ability to form biofilms.
Results: We deleted SMU.1655c to create S. mutans MMZ1129. While there was no change in ser gene expression, MMZ1129 displayed minor defects in growth and competence development, but an increased ability to form biofilms. Interestingly, RNA-Seq revealed increased transcription of the mub operon, loss of expression of two TetR-family regulators (mubR and SMU.1361c), and a 9-kbp deletion within TnSmu2 (SMU. 1348c-1365c). Although MubR is known to positively regulate the mub operon, strain MMZ1129 had enhanced expression of mub genes, which we found was due to enhanced transcription of SMU.1367c. Subsequently, we found that deletion of SMU.1361c led to a 100-fold increase in SMU.1367c mRNA, which in turn led to higher mub gene expression. Similar deletions were not found in other SMU.1655c mutants, so no cause-and-effect relationship exists between deletion of SMU.1655c and loss of TnSmu2 sequences. However, a mutant (1655cCCSS) in which a double-cysteine motif in SMU.1655c was replaced with serines displayed reduced expression of mub genes.
Conclusions: This study has identified a new regulator of TnSmu2 gene expression and that a small peptide, possibly involved in redox sensing, can influence transcription of genes in this genomic island.