Identification of pel , a Streptococcus pyogenes Locus That Affects both Surface and Secreted Proteins

Abstract

A Tn917 insertion mutant of an M49 serotype, opacity factor-positive Streptococcus pyogenes, was isolated. It had the following phenotypes: decreased β-hemolysis mediated by streptolysin S, reduction in the activity of a secreted cysteine protease and streptokinase, and an altered immunoglobulin and fibrinogen-binding phenotype. The site of insertion of Tn917 into the chromosome and the surrounding sequence, thepel region (pleiotropic effect locus), was determined. Phage A25 transduction confirmed that the pleiotropic changes in phenotype could be cotransduced with Tn917. Thepel region was cloned and sequenced, and the transposon was found to be inserted upstream of a single open reading frame which led to a failure to transcribe a 500-base mRNA. The loss of this transcript decreased the transcription of emm and speBgenes and reduced the secretion of streptokinase. Enhanced Pel expression from a nisin-inducible plasmid resulted in increased message levels for emm in a wild-type organism. Characterization of the pel mutant provides evidence for the coordinated regulation of secreted and surface proteins and suggests the existence of a new global regulatory factor in S. pyogenes.

Mutational analysis of the group A streptococcal operon encoding streptolysin S and its virulence role in invasive infection

“…Earlier studies establishing the role of the sag locus in SLS production employed transposon mutants (Betschel et al ., 1998;Li et al ., 1999) or insertional inactivation mutants (Nizet et al ., 2000;Biswas et al ., 2001) likely to exert polar effects on transcription of downstream genes in the operon. To assess the specific requirement of individual sag genes in SLS production, we combined precise in-frame allelic exchange mutagenesis with single-gene complementation analysis.…”

Synthesis of group A streptococcal virulence factors is controlled by a regulatory RNA molecule

“…Total RNA was prepared according to a previously described protocol (Li et al, 1999). In brief, total RNA was prepared from samples equalized to the same number of cells.…”

“…Recently, the analysis of a Tn 917 transposon library of an M49 GAS strain revealed a gene locus, p leiotropic e ffect l ocus ( pel ), with a pleiotropic effect on the expression of a number of streptococcal exoproteins (Li et al ., 1999). The pel locus comprises sagA , which is the structural gene for SLS (Nizet et al ., 2000).…”

“…The genes located downstream of the sagA gene, sagB to sagI , have been shown to be required for the proper secretion of SLS, because they are involved in processing and export of SLS. As described, the pel locus exerts its effects on virulence factors, including M-protein, SpeB and Ska (Li et al ., 1999;Biswas et al ., 2001). Another report showed that in vivo passage of a pel -deficient mutant selected for mutants with restored production of SLS and M-protein but not SpeB or Ska (Eberhard et al ., 2001).…”

Comparative genome‐wide analysis of small RNAs of major Gram‐positive pathogens: from identification to application

“…A first GAS sRNA sagA was discovered during the identification and functional characterization of the major haemolysin of GAS, streptolysin S (SLS/SagA) (Li et al, 1999;Carr et al, 2001). Mangold and colleagues later reported that the mRNA transcribed from the sls gene most likely acts as sRNA and effectively regulates expression of emm (M protein), sic (streptococcal inhibitor of complement) and nga (NAD-glycohydrolase) at the transcriptional level, and expression of SpeB (cystein protease) at the post-transcriptional level (Mangold et al, 2004).…”