Human group V secretory phospholipase A2 is associated with lipid rafts and internalized in a flotillin‑dependent...

The mechanisms of secretory phospholipase A2 (sPLA2) action are not understood clearly. Previously, it was suggested that sPLA2s are internalized into cells for the targeting of sPLA2 to intracellular action sites. However, the mechanisms for sPLA2 internalization remain to be identified. The present study demonstrated for the first time that human group V sPLA2 (hVPLA2) is associated with lipid rafts and is internalized in a flotillin‑dependent pathway. The lipid raft association was probed by cholesterol‑sensitive enrichment of hVPLA2 in low‑density fractions and co‑patching of ganglioside GM1 rafts through cross‑linking of hVPLA2 in HEK293 and CHO cells. The hVPLA2 associated with lipid rafts was shown to be internalized into HEK293 cells at a relatively rapid rate (t1/2 =16 min) and this internalization was inhibited by the knockdown of flotillin‑1, but not by chlorpromazine, an inhibitor of clathrin‑mediated endocytosis. Moreover, internalized hVPLA2 was shown to be colocalized extensively with flotillin‑1 in a punctate structure, but not caveolin‑1. These data revealed that the internalization of hVPLA2 is mediated by flotillin‑1. Attenuation of arachidonic acid release from plasma membrane through the association of hVPLA2 with lipid rafts suggested that this association with lipid rafts may be important in protecting mammalian cells from excessive degradation of plasma membrane and trafficking hVPLA2 into intracellular targets.