SPE B-induced apoptosis in A549 cells is mediated through _v3 integrin and Fas

Institute of Basic Medical Sciences, Departments of Biochemistry, Microbiology and Immunology, Medical Laboratory Science and Biotechnology, and Pediatrics, National Cheng Kung University Medical College, Tainan, Taiwan; National Applied Research Laboratories, National Laboratory Animal Center, Taipei, Taiwan; Institute of Medical Sciences, School of Medicine, Tzu Chi University, Hualien, Taiwan

^* To whom correspondence should be addressed. Email: lin1223{at}mail.tcu.edu.tw.

	Abstract

Our previous work suggested that streptococcal pyrogenic exotoxin B (SPE B)-induced apoptosis is mediated through a receptor-like mechanism. In this study, we have identified _v3 and Fas as the SPE B receptors for this function. The SPE B fragment without the RGD motif and G308S, a SPE B mutant with the RSD motif, induced less apoptosis than did native SPE B, suggesting that the RGD motif is critical for SPE B-induced apoptosis. FITC-SPE B binding assays and immunoprecipitation analysis showed that SPE B specifically interacted with _v3. Anti-_v3 antibody partially inhibited SPE B-induced apoptosis but had no effect on G308S-induced apoptosis. In addition, the Fas binding to SPE B was verified in an affinity column and an immunoprecipitation analysis. Anti-Fas antibody inhibited SPE B- and G308S-induced apoptosis in a dose dependent manner, suggesting that Fas-mediated SPE B-induced apoptosis also occurs RGD-independently. Both anti-_v3 and anti-Fas antibodies synergistically inhibited SPE B-induced apoptosis. The apoptotic cascades were activated by SPE B and G308S, with a little delay by the latter. After SPE B binding, the cell surface level of _v3 but not of Fas was decreased. The decreased _v3 level was restored by proteasome inhibitor MG132 treatment, suggesting a SPE B-mediated endocytosis of integrin _v3 via the ubiquitin-proteasome system. Taken together, our results demonstrate that the SPE B-induced apoptosis is mediated through _v3 integrin and Fas in a synergistic manner.