An invertebrate β-integrin mediates coelomocyte phagocytosis via activation of septin2 and 7 but not septin10

Abstract

We have previously confirmed that β-integrin from Apostichopus japonicus (designated AjITGB) binds LPS and mediates the immune response. In this study, we found that AjITGB positively promoted the echinoderm immune cells located in the coelomic cavity (designated as coelomocyte) phagocytic activities. Flow cytometry assay indicated that the phagocytic percentage was significantly decreased, by a 37.3% and 41.36%, after AjITGB siRNA inference in vitro and in vivo, respectively, a result consistent with the decrease observed with an anti-AjITGB antibody blocking treatment. These decreased phagocytic activities were partially recovered by rAjITGB supplementation. To better understand the molecular mechanism underlying this phagocytosis, three phagocytic-related proteins, including Ajseptin2, Ajseptin7 and Ajseptin10, were cloned and further characterized. Completely consistent expression profiles were detected between AjITGB and Ajseptin2/7, at both the mRNA and protein levels, in V. splendidus-challenged sea cucumber. Furthermore, Ajseptin2/7 expression levels were significantly down-regulated in the AjITGB silencing group and could be partially recovered to its original level after rAjITGB administration. However, Ajseptin10 displayed no significant change in the same condition. A phagocytic assay further indicated that Ajseptin2/7, but not Ajseptin10, was crucial in mediating coelomocyte phagocytosis. The knockdown of the three genes by specific siRNAs indicated that Ajseptin2/7 significantly decreased coelomocyte phagocytosis, and this decrease was completely recovered after rAjITGB supplementation. There was no significant change in the phagocytosis rate in both the AjSEPT10 siRNA interference and rAjITGB supplementation groups. All our results confirmed that AjITGB modulates coelomocyte phagocytosis via the activation of Ajseptin2/7 but not AjSEPT10 and further supported the divergent roles of Ajseptins in AjITGB-mediated phagocytosis.

Keywords: Apostichopus japonicas; Flow cytometry; Integrins; Phagocytosis; Septins.