Roles of PknB and CslA in cell wall morphogenesis ofStreptomyces

Abstract Streptomyces are filamentous bacteria living mostly in the soil and best known for their ability to produce specialised metabolites, including antibiotics. They grow as tip-extending, branching hyphal filaments to form a multicellular mycelium. New branches are established by the formation of a new growth zone on the lateral cell wall. Proteins responsible for this process are organised in complexes called polarisomes, with DivIVA being the best described component. We have developed a genetic screen based on DivIVA overexpression to identify proteins potentially interacting with DivIVA in Streptomyces albus . Among the hits was the morphology controlling protein rodA and the kinase pknB . We confirmed that deletion of pknB rescues S. albus from the effects of overexpressing DivIVA. Next, we studied the impact of pknB deletion in Streptomyces. A mass spectrometry phosphoproteome analysis indicated that absence of PknB alters the phosphorylation state of CslA, a cellulose synthase-like protein, containing a glycosyltransferase domain. We demonstrate that a phosphoablative mutant of CslA impacts its activity in β-glucan synthesis, as displayed by a hypersensitivity to lysozyme. Our findings highlight the role of PknB in maintaining the cellular structure and integrity of the Streptomyces cell wall. Importance A thorough understanding of the regulatory mechanisms of cell wall biosynthesis and integrity in Streptomyces is fundamentally important, as these are linked to antibiotic production. This study investigates the role of the kinase PknB, which we identified via a genetic screen. Through a phosphoproteome analysis, we identified that PknB regulates the activity of CslA, a protein crucial for β-glucan synthesis. By demonstrating the impact of PknB, this research provides novel insights into the molecular pathways influencing Streptomyces resilience. This research not only enhances our understanding of cell morphogenesis but also has potential implications for developing new strategies to optimize antibiotic production.