ABSTRACT
Streptococcus pyogenes (group A streptococcus (GAS)) is a pathogen that invades non-phagocytic host cells, and causes a variety of acute infections such as pharyngitis. Our group previously reported that intracellular GAS is effectively degraded by the host-cell autophagic machinery, and that a cholesterol-dependent cytolysin, streptolysin O (SLO), is associated with bacterial escape from endosomes in epithelial cells. However, the details of both the intracellular behavior of GAS and the process leading to its autophagic degradation remain unknown. In this study, we found that two host small G proteins, Rab5 and Rab7, were associated with the pathway of autophagosome formation and the fate of intracellular GAS. Rab5 was involved in bacterial invasion and endosome fusion. Rab7 was clearly multifunctional, with roles in bacterial invasion, endosome maturation, and autophagosome formation. In addition, this study showed that the bacterial cytolysin SLO supported the escape of GAS into the cytoplasm from endosomes, and surprisingly, a SLO-deficient mutant of GAS was viable longer than the wild-type strain although it failed to escape the endosomes. This intracellular behavior of GAS is unique and distinct from that of other types of bacterial invaders. Our results provide a new picture of GAS infection and host-cell responses in epithelial cells.
Subject(s)
Autophagy , Intracellular Space/microbiology , Streptolysins/metabolism , rab GTP-Binding Proteins/metabolism , rab5 GTP-Binding Proteins/metabolism , Bacterial Proteins/metabolism , Biomarkers/metabolism , Endosomes/microbiology , Endosomes/ultrastructure , HeLa Cells , Humans , Intracellular Space/metabolism , Lysosomes/microbiology , Lysosomes/ultrastructure , Microbial Viability , Models, Biological , Mutant Proteins/metabolism , Phagosomes/metabolism , Phagosomes/microbiology , Phagosomes/ultrastructure , Protein Transport , Streptococcus pyogenes/cytology , Streptococcus pyogenes/metabolism , Streptococcus pyogenes/ultrastructure , Time Factors , rab7 GTP-Binding ProteinsABSTRACT
BACKGROUND: A supernumerary tooth is an extra tooth above the normal number, of which approximately 90% occur in the premaxillary region and show rudimentary forms of crown morphology. Most cases occur singly, with bilateral occurrence in the maxillary canine regions very rare in children with no other associated diseases or syndromes. CASE REPORT: A case of a 14-year-old boy with bilateral supernumerary teeth with normal crown shapes in both the maxillary canine and mandibular premolar regions. The supernumerary teeth in the maxilla were diagnosed at 12.2 years of age and in the mandible at 14.1 years of age. CONCLUSION: Four supernumerary teeth in the maxillary and mandibular canine-premolar region is a rare finding.
Subject(s)
Tooth, Supernumerary , Adolescent , Age Determination by Teeth , Bicuspid , Cuspid , Dentition, Permanent , Humans , Male , Mandible , MaxillaABSTRACT
Helicobacter pylori vacuolating cytotoxin, VacA, which causes vacuolation of gastric epithelial cells and other types of cultured cells, is known to stimulate apoptosis via a mitochondria-dependent pathway. In the present study, we examined the mechanisms of VacA-induced mitochondrial damage. Intracellular VacA localization was monitored by immunostaining and confocal microscopy; in AZ-521 cells in which cytochrome c release was stimulated, most of VacA was localized to vacuoles rather than mitochondria. VacA reduced the membrane potential of isolated mitochondria without inducing cytochrome c release, suggesting that it did not act directly to induce cytochrome c release from mitochondria and that in intact cells, VacA-induced cytochrome c release involved apoptosis-related factor(s), such as a proapoptotic Bcl-2 family protein. In agreement, flow cyto-metric analyses using antibodies specific for activated Bax revealed that intracellular Bax was activated by VacA in a concentration- and time-dependent manner. Using active form-specific antibodies, we also observed that the Bcl-2 family protein, Bak, was activated. By confocal microscopy, Bax and Bak were activated in AZ-521 cells in which cyto-chrome c release was induced by VacA. In addition, small interfering RNA-induced silencing of the bax gene resulted in reduction of VacA-stimulated cytochrome c release, consistent with a contribution of VacA-induced Bax activation to cytochrome c release. NH4Cl enhanced both VacA-induced vacuolation and Bax activation, whereas Bax activation was not inhibited by bafilomycin A1, which inhibited vacuolation caused by VacA. These results suggest that VacA acts through different signaling pathways to induce apoptosis via Bax activation, independent of vacuolation.
Subject(s)
Bacterial Proteins/physiology , Cytochromes c/metabolism , Helicobacter pylori/metabolism , bcl-2 Homologous Antagonist-Killer Protein/metabolism , bcl-2-Associated X Protein/metabolism , Apoptosis , Bacterial Proteins/chemistry , Cell Death , Cell Line , Cell Line, Tumor , Flow Cytometry , HeLa Cells , Humans , Immunohistochemistry , Macrolides/pharmacology , Microscopy, Confocal , Microscopy, Fluorescence , Mitochondria/metabolism , Protons , RNA, Small Interfering/metabolism , Time Factors , Transfection , Vacuoles/metabolismABSTRACT
We found that the autophagic machinery could effectively eliminate pathogenic group A Streptococcus (GAS) within nonphagocytic cells. After escaping from endosomes into the cytoplasm, GAS became enveloped by autophagosome-like compartments and were killed upon fusion of these compartments with lysosomes. In autophagy-deficient Atg5-/- cells, GAS survived, multiplied, and were released from the cells. Thus, the autophagic machinery can act as an innate defense system against invading pathogens.
