Pigpen, a nuclear coiled body component protein, is involved in angiogenesis

We previously reported that puromycin-insensitive leucyl-specific aminopeptidase (PILSAP) is required for vascular endothelial growth factor (VEGF)- and basic fibroblast growth factor (bFGF)-induced angiogenesis and for endothelial differentiation from embryonic stem (ES) cells via the aminopeptidase activity of PILSAP. In this study, we searched for molecules that function during angiogenesis with PILSAP. We performed proteome analysis of nuclear extracts from embryoid bodies (EBs) made from ES cells transfected with mutant PILSAP lacking aminopeptidase activity and mock EBs. We identified pigpen, a 67-kDa nuclear coiled body component protein. Immunoprecipitation and western blotting demonstrated the binding of PILSAP and pigpen in endothelial cells (ECs), and this interaction was enhanced by VEGF and bFGF. Pigpen was reported to be expressed in actively growing ECs such as those in embryos and tumors. However, whether Pigpen is involved in angiogenesis is not known. Therefore, we examined the effect of pigpen on angiogenesis by silencing pigpen with siRNA (siPigpen). Compared with scrambled RNA (scrPigpen), transfection of siPigpen into mouse ECs inhibited proliferation, migration, and network formation. These results were confirmed with other two siRNAs. Moreover, siPigpen suppressed bFGF-induced angiogenesis in a Matrigel plug assay, and injection of siPigpen into Lewis lung carcinoma cell tumors implanted subcutaneously into 5-week-old C57/BL male mice prevented tumor growth and tumor angiogenesis. These data indicate that Pigpen is involved in angiogenesis and that pigpen may be a target for blocking tumor angiogenesis.