The StkP/PhpP Signaling Couple in Streptococcus pneumoniae: Cellular Organization and Physiological Characterization
Abstract
The StkP/PhpP Signaling Couple in Streptococcus pneumoniae : Cellular Organization and Physiological Characterization ▿ † Makoto Osaki 1 , 2 , Tania Arcondéguy 1 , Amandine Bastide 1 , Christian Touriol 1 , Hervé Prats 1 , and Marie-Claude Trombe 1 , * 1 INSERM, U858, Toulouse, F-31432 France, and IFR31 Université Toulouse III Paul-Sabatier, Toulouse, France 2 National Institute of Animal Health, National Agriculture and Food Research Organization, Tsukuba, Ibaraki 305-0856, Japan ABSTRACT In Streptococcus pneumoniae , stkP and phpP , encoding the eukaryotic-type serine-threonine kinase and PP2C phosphatase, respectively, form an operon. PhpP has the features of a so-called “soluble” protein, whereas StkP protein is membrane associated. Here we provide the first genetic and physiological evidence that PhpP and StkP, with antagonist enzymatic activities, constitute a signaling couple. The StkP-PhpP couple signals competence upstream of the competence-specific histidine kinase ComD, receptor for the oligopeptide pheromone “competence stimulating peptide.” We show that PhpP activity is essential in a stkP + genetic background, suggesting tight control of StkP activity by PhpP. Proteins PhpP and StkP colocalized to the cell membrane subcellular fraction and likely belong to the same complex, as revealed by coimmunoprecipitation in cellular extracts. Specific coimmunoprecipitation of the N-kinase domain of StkP and PhpP recombinant proteins by PhpP-specific antibodies demonstrates direct interaction between these proteins. Consistently, flow cytometry analysis allowed the determination of the cytoplasmic localization of PhpP and of the N-terminal kinase domain of StkP, in contrast to the periplasmic localization of the StkP C-terminal PASTA ( p enicillin-binding protein a nd s erine- t hreonine kinase a ssociated) domain. A signaling route involving interplay between serine, threonine, and histidine phosphorylation is thus described for the first time in this human pathogen.