title: Protochlamydia induces apoptosis of human HEp-2 cells through mitochondrial dysfunction mediated by chlamydial protease-like activity factor.
abstract: Obligate amoebal endosymbiotic bacterium Protochlamydia with ancestral pathogenic chlamydial features evolved to survive within protist hosts, such as Acanthamoba, 0.7-1.4 billion years ago, but not within vertebrates including humans. This observation raises the possibility that interactions between Protochlamydia and human cells may result in a novel cytopathic effect, leading to new insights into host-parasite relationships. Previously, we reported that Protochlamydia induces apoptosis of the immortalized human cell line, HEp-2. In this study, we attempted to elucidate the molecular mechanism underlying this apoptosis. We first confirmed that, upon stimulation with the bacteria, poly (ADP-ribose) polymerase (PARP) was cleaved at an early stage in HEp-2 cells, which was dependent on the amount of bacteria. A pan-caspase inhibitor and both caspase-3 and -9 inhibitors similarly inhibited the apoptosis of HEp-2 cells. A decrease of the mitochondrial membrane potential was also confirmed. Furthermore, lactacystin, an inhibitor of chlamydial protease-like activity factor (CPAF), blocked the apoptosis. Cytochalasin D also inhibited the apoptosis, which was dependent on the drug concentration, indicating that bacterial entry into cells was required to induce apoptosis. Interestingly, Yersinia type III inhibitors (ME0052, ME0053, and ME0054) did not have any effect on the apoptosis. We also confirmed that the Protochlamydia used in this study possessed a homologue of the cpaf gene and that two critical residues, histidine-101 and serine-499 of C. trachomatis CPAF in the active center, were conserved. Thus, our results indicate that after entry, Protochlamydia-secreted CPAF induces mitochondrial dysfunction with a decrease of the membrane potential, followed by caspase-9, caspase-3 and PARP cleavages for apoptosis. More interestingly, because C. trachomatis infection can block the apoptosis, our finding implies unique features of CPAF between pathogenic and primitive chlamydiae.
authors: Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, Sapporo, Hokkaido, Japan.
authors: Junji
authors: J
authors: Matsuo
authors: Shinji
authors: S
authors: Nakamura
authors: Atsushi
authors: A
authors: Ito
authors: Tomohiro
authors: T
authors: Yamazaki
authors: Kasumi
authors: K
authors: Ishida
authors: Yasuhiro
authors: Y
authors: Hayashi
authors: Mitsutaka
authors: M
authors: Yoshida
authors: Kaori
authors: K
authors: Takahashi
authors: Tsuyoshi
authors: T
authors: Sekizuka
authors: Fumihiko
authors: F
authors: Takeuchi
authors: Makoto
authors: M
authors: Kuroda
authors: Hiroki
authors: H
authors: Nagai
authors: Kyoko
authors: K
authors: Hayashida
authors: Chihiro
authors: C
authors: Sugimoto
authors: Hiroyuki
authors: H
authors: Yamaguchi
mesh_terms: D000595:Amino Acid Sequence; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002460:Cell Line; D002689:Chlamydia / Q000235:genetics / Q000378:metabolism*; D010450:Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008928:Mitochondria / Q000378:metabolism*; D008969:Molecular Sequence Data; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D059748:Proteolysis; D016415:Sequence Alignment; D037521:Virulence Factors / Q000378:metabolism
chemical_list: D037521:Virulence Factors; D011065:Poly(ADP-ribose) Polymerases; D010450:Endopeptidases; D053148:Caspase 3; D053453:Caspase 9; C425923:CPA factor