title: Endocytosis of Streptococcus pneumoniae via the polymeric immunoglobulin receptor of epithelial cells relies on clathrin and caveolin dependent mechanisms.
abstract: Colonization of Streptococcus pneumoniae (pneumococci) is a prerequisite for bacterial dissemination and their capability to enter the bloodstream. Pneumococci have evolved various successful strategies to colonize the mucosal epithelial barrier of humans. A pivotal mechanism of host cell invasion implicated with invasive diseases is promoted by the interaction of pneumococcal PspC with the polymeric Ig-receptor (pIgR). However, the mechanism(s) of pneumococcal endocytosis and the intracellular route of pneumococci upon uptake by the PspC-pIgR-interaction are not known. Here, we demonstrate by using a combination of pharmacological inhibitors and genetics interference approaches the involvement of active dynamin-dependent caveolae and clathrin-coated vesicles for pneumococcal uptake via the PspC-pIgR mechanism. Depleting cholesterol from host cell membranes and disruption of lipid microdomains impaired pneumococcal internalization. Moreover, chemical inhibition of clathrin or functional inactivation of dynamin, caveolae or clathrin by RNA interference significantly affected pneumococcal internalization suggesting that clathrin-mediated endocytosis (CME) and caveolae are involved in the bacterial uptake process. Confocal fluorescence microscopy of pIgR-expressing epithelial cells infected with pneumococci or heterologous Lactococcus lactis expressing PspC demonstrated bacterial co-localization with fluorescent-tagged clathrin and early as well as recycling or late endosomal markers such as Lamp1, Rab5, Rab4, and Rab7, respectively. In conclusion these data suggest that PspC-promoted uptake is mediated by both CME and caveolae. After endocytosis pneumococci are routed via the endocytic pathway into early endosomes and are then sorted into recycling or late endosomes, which can result in pneumococcal killing in phagolysosomes or transcytosis via recycling endosomes.
authors.affiliation: Department Genetics of Microorganisms, Interfaculty Institute for Genetics and Functional Genomics, Ernst-Moritz-Arndt Universität Greifswald, Friedrich-Ludwig-Jahn-Strasse 15a, D-17487 Greifswald, Germany; Pediatric Infectious Diseases, Department of Pediatrics, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.
authors.forename: Tauseef M
authors.initials: TM
authors.lastname: Asmat
authors.affiliation: Department Genetics of Microorganisms, Interfaculty Institute for Genetics and Functional Genomics, Ernst-Moritz-Arndt Universität Greifswald, Friedrich-Ludwig-Jahn-Strasse 15a, D-17487 Greifswald, Germany; Department of Laboratory Medicine, Medical Protein Chemistry, Malmö University Hospital, Lund University, Malmö, Sweden.
authors.forename: Vaibhav
authors.initials: V
authors.lastname: Agarwal
authors.affiliation: Department Genetics of Microorganisms, Interfaculty Institute for Genetics and Functional Genomics, Ernst-Moritz-Arndt Universität Greifswald, Friedrich-Ludwig-Jahn-Strasse 15a, D-17487 Greifswald, Germany.
authors.forename: Malek
authors.initials: M
authors.lastname: Saleh
authors.affiliation: Department Genetics of Microorganisms, Interfaculty Institute for Genetics and Functional Genomics, Ernst-Moritz-Arndt Universität Greifswald, Friedrich-Ludwig-Jahn-Strasse 15a, D-17487 Greifswald, Germany. Electronic address: sven.hammerschmidt@uni-greifswald.de.
authors.forename: Sven
authors.initials: S
authors.lastname: Hammerschmidt
mesh_terms: D000818:Animals; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000378:metabolism; D022461:Caveolins / Q000378:metabolism*; D002460:Cell Line; D002966:Clathrin / Q000378:metabolism*; D004285:Dogs; D004705:Endocytosis; D004847:Epithelial Cells / Q000382:microbiology / Q000502:physiology*; D006801:Humans; D011485:Protein Binding; D019056:Receptors, Polymeric Immunoglobulin / Q000378:metabolism*; D013296:Streptococcus pneumoniae / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D022461:Caveolins; D002966:Clathrin; D019056:Receptors, Polymeric Immunoglobulin; C110344:SpsA protein, Streptococcus pneumoniae
keywords: Adherence; Caveolin; Clathrin; Dynamin; Endocytosis; Polymeric Ig receptor; PspC; Rab; Streptococcus pneumoniae