title: Knockdown of legumain inhibits cleavage of annexin A2 in the mouse kidney.
abstract: Legumain (EC 3.4.22.34) is an asparaginyl endopeptidase. Strong legumain activity was observed in the mouse kidney, and legumain was highly expressed in tumors. We previously reported that bovine kidney annexin A2 was co-purified with legumain and that legumain cleaved the N-terminal region of annexin A2 at an Asn residue in vitro. In this study, to determine whether annexin A2 is cleaved by legumain in vivo, siRNA-lipoplex targeting mouse legumain was injected into mouse tail veins. Mouse kidneys were then isolated and the effect of knockdown of legumain expression on annexin A2 cleavage was examined. The results showed that both legumain mRNA and protein expression levels were decreased in the siRNA-treated mouse kidneys and that legumain activity toward a synthetic substrate, Z-Ala-Ala-Asn-MCA, was decreased by about 40% in the kidney but not in the liver or spleen. Furthermore, cleavage of annexin A2 at the N-terminal region was decreased in the mouse kidney that had been treated with the legumain siRNA-lipoplex. These results suggest that legumain siRNA was delivered to the kidney by using LipoTrust and that the reduced legumain expression inhibited legumain-induced degradation of annexin A2 in vivo.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita-ku, Sapporo 060-0812, Japan. t-yamane@pharm.hokudai.ac.jp
authors.forename: Takuya
authors.initials: T
authors.lastname: Yamane
authors.forename: Rei
authors.initials: R
authors.lastname: Hachisu
authors.forename: Motoki
authors.initials: M
authors.lastname: Yuguchi
authors.forename: Keisuke
authors.initials: K
authors.lastname: Takeuchi
authors.forename: Sato
authors.initials: S
authors.lastname: Murao
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Yamamoto
authors.forename: Hisakazu
authors.initials: H
authors.lastname: Ogita
authors.forename: Toshihide
authors.initials: T
authors.lastname: Takasawa
authors.forename: Iwao
authors.initials: I
authors.lastname: Ohkubo
authors.forename: Hiroyoshi
authors.initials: H
authors.lastname: Ariga
mesh_terms: D000818:Animals; D017306:Annexin A2 / Q000378:metabolism*; D001216:Asparagine / Q000378:metabolism; D002417:Cattle; D003546:Cysteine Endopeptidases / Q000235:genetics* / Q000378:metabolism; D055785:Gene Knockdown Techniques / Q000379:methods*; D007668:Kidney / Q000201:enzymology*; D051379:Mice; D008810:Mice, Inbred C57BL; D041681:NIH 3T3 Cells; D059748:Proteolysis; D034741:RNA, Small Interfering / Q000235:genetics*; D014162:Transfection / Q000379:methods; D000099240:Asparaginyl Endopeptidase
chemical_list: D017306:Annexin A2; D001216:Asparagine; D003546:Cysteine Endopeptidases; D034741:RNA, Small Interfering; D000099240:Asparaginyl Endopeptidase