Inactivation of proprotein convertase, PACE4, by α1-antitrypsin Portland (α1-PDX), a blocker of proteolytic activation of bone morphogenetic protein during embryogenesis:: Evidence that PACE4 is able to form an SDS-stable acyl intermediate with α1-PDX

被引:44
作者
Tsuji, A [1 ]
Hashimoto, E [1 ]
Ikoma, T [1 ]
Taniguchi, T [1 ]
Mori, K [1 ]
Nagahama, M [1 ]
Matsuda, Y [1 ]
机构
[1] Univ Tokushima, Fac Engn, Dept Biol Sci & Technol, Tokushima 7708506, Japan
关键词
alpha; 1-antitrypsin; BMP processing; PACE4; 1-PDX; SPC;
D O I
10.1093/oxfordjournals.jbchem.a022491
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
PACE4 (SPC4), a member of the subtilisin-like proprotein convertase (SPC) family of proteases that cleave at paired basic amino acids, exhibits a dynamic expression pattern during embryogenesis and colocalizes with bone morphogenetic proteins (BMPs), Recently Cui et at. reported that the ectopic expression of alpha 1-antitrypsin variant Portland (alpha 1-PDX), an engineered serpin that contains the minimal SPC consensus motif in its reactive loop, blocks the proteolytic activation of BMP4, leading to abnormal embryogenic development [Cui, Y, et at (1998) EMBO J, 17, 4735-4743], TGF beta-related factors such as BMPs are synthesized as inactive precursors and activated by limited proteolysis at multibasic amino acids. Therefore, an alpha 1-PDX-inhibitable protease is thought to participate in BMP activation, However, conflicting properties, including sensitivity to alpha 1-PDX, have been reported for PACE4, In this study, we examined whether alpha 1-PDX is responsible for the inhibition of PACE4 by measuring the protease/inhibitor complex directly. Here we show that alpha 1-PDX has the ability to form an SDS-stable acyl-intermediate (180 kDa) with PACE4 in vivo and in vitro. Further, we characterized the PACE4 secreted into the culture medium from Cos-1 cells by a specific immunological assay. An alpha 1-PDX-insensitive and decanoyl-RVKR-chloromethylketone-sensitive 60-kDa protease(s) is greatly activated in conditioned medium by PACE4 overexpression, suggesting that the activation of an unknown protease(s) other than PACE4 is the cause of the variation in the properties of PACE4, PACE4 is a Ca2+-dependent protease with an optimal Ca2+ requirement of 2 mM, and shows its highest activity at weakly basic pH, PACE4 activity is completely inhibited by EDTA and EGTA, but not by leupeptin, These results show that PACE4 activity can be inhibited by alpha 1-PDX as well as furin (SPC1) and suggest that the inhibition of PACE4-mediated activation of factors such as BMPs by (alpha 1-PDX causes abnormal embryogenic development.
引用
收藏
页码:591 / 603
页数:13
相关论文
共 51 条
[1]   Developmental expression of a novel Kexin family protease, PACE4E, in the rat olfactory system [J].
Akamatsu, T ;
Daikoku, S ;
Nagamune, H ;
Yoshida, S ;
Mori, K ;
Tsuji, A ;
Matsuda, Y .
HISTOCHEMISTRY AND CELL BIOLOGY, 1997, 108 (02) :95-103
[2]  
AZAYAN AV, 1995, J BIOL CHEM, V270, P8201
[3]  
BEAUBIEN G, 1995, CELL TISSUE RES, V279, P539
[4]   alpha 1-antitrypsin portland inhibits processing of precursors mediated by proprotein convertases primarily within the constitutive secretory pathway [J].
Benjannet, S ;
Savaria, D ;
Laslop, A ;
Munzer, JS ;
Chretien, M ;
Marcinkiewicz, M ;
Seidah, NG .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1997, 272 (42) :26210-26218
[5]  
BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3
[6]   COMPARATIVE PROTEOLYTIC PROCESSING OF RAT PROSOMATOSTATIN BY THE CONVERTASES PC1, PC2, FURIN, PACE4 AND PC5 IN CONSTITUTIVE AND REGULATED SECRETORY PATHWAYS [J].
BRAKCH, N ;
GALANOPOULOU, AS ;
PATEL, YC ;
BOILEAU, G ;
SEIDAH, NG .
FEBS LETTERS, 1995, 362 (02) :143-146
[7]   PC8, a new member of the convertase family [J].
Bruzzaniti, A ;
Goodge, K ;
Jay, P ;
Taviaux, SA ;
Lam, MHC ;
Berta, P ;
Martin, TJ ;
Moseley, JM ;
Gillespie, MT .
BIOCHEMICAL JOURNAL, 1996, 314 :727-731
[8]   SPC4, SPC6, and the novel protease SPC7 are coexpressed with bone morphogenetic proteins at distinct sites during embryogenesis [J].
Constam, DB ;
Calfon, M ;
Robertson, JJ .
JOURNAL OF CELL BIOLOGY, 1996, 134 (01) :181-191
[9]   Regulation of bone morphogenetic protein activity by pro domains and proprotein convertases [J].
Constam, DB ;
Robertson, EJ .
JOURNAL OF CELL BIOLOGY, 1999, 144 (01) :139-149
[10]   BMP-4 is proteolytically activated by furin and/or PC6 during vertebrate embryonic development [J].
Cui, YZ ;
Jean, F ;
Thomas, G ;
Christian, JL .
EMBO JOURNAL, 1998, 17 (16) :4735-4743