Sato T, Kondo T, Fujisawa T, Seiki M, Ito A
Department of Biochemistry, School of Pharmacy, Tokyo University of Pharmacy and Life Science, Horinouchi, Hachioji, Tokyo 192-0392, Japan.
J Biol Chem. 1999 Dec 24;274(52):37280-4. doi: 10.1074/jbc.274.52.37280.
We investigated the gene expression and intracellular activity of processing protease furin and its involvement in the process of membrane type 1-matrix metalloproteinase (MT1-MMP) activation in rabbit dermal fibroblasts. When the rabbit fibroblasts were treated with concanavalin A (ConA), pro-MMP-2 was converted to an active 62-kDa MMP-2 through the appearance of a 64-kDa intermediate MMP-2. The ConA-induced pro-MMP-2 activation resulted from increasing the gene expression and production of MT1-MMP in the rabbit fibroblasts. Reverse transcriptase-polymerase chain reaction demonstrated that in rabbit dermal fibroblasts furin mRNA was detected and, unlike MT1-MMP, was not increased by ConA. These findings are further supported by the fact that the intracellular furin activity also was constitutively detected and was unchanged by the ConA treatment. Very similar phenomena were also observed in human uterine cervical fibroblasts, which are known to produce MT1-MMP by ConA stimulation. These results suggest that the expression of the furin gene and the intracellular activity are not regulated by ConA. On the other hand, neither a synthetic furin inhibitor, decanoyl-RVKR-CH(2)Cl (25-100 microM) nor a furin antisense oligonucleotide (40 microM) inhibited the MT1-MMP-mediated pro-MMP-2 activation in ConA-treated rabbit dermal fibroblasts, whereas these compounds interfered with pro-MMP-2 activation in ConA-treated human uterine cervical fibroblasts. Nonetheless, the furin antisense oligonucleotide completely suppressed furin gene expression in both rabbit and human fibroblasts. These results suggest that furin does not participate in the process of MT1-MMP activation induced by ConA in rabbit dermal fibroblasts.
我们研究了加工蛋白酶弗林蛋白酶的基因表达和细胞内活性，及其在兔真皮成纤维细胞中膜型1基质金属蛋白酶（MT1-MMP）激活过程中的作用。当兔成纤维细胞用伴刀豆球蛋白A（ConA）处理时，前MMP-2通过64 kDa中间MMP-2的出现转化为活性62 kDa的MMP-2。ConA诱导的前MMP-2激活是由于兔成纤维细胞中MT1-MMP的基因表达和产生增加。逆转录聚合酶链反应表明，在兔真皮成纤维细胞中检测到弗林蛋白酶mRNA，与MT1-MMP不同，它不会被ConA增加。细胞内弗林蛋白酶活性也被持续检测到且不受ConA处理影响这一事实进一步支持了这些发现。在已知通过ConA刺激产生MT1-MMP的人子宫颈成纤维细胞中也观察到了非常相似的现象。这些结果表明弗林蛋白酶基因的表达和细胞内活性不受ConA调节。另一方面，合成的弗林蛋白酶抑制剂癸酰-RVKR-CH(2)Cl（25 - 100 microM）和弗林蛋白酶反义寡核苷酸（40 microM）均未抑制ConA处理的兔真皮成纤维细胞中MT1-MMP介导的前MMP-2激活，而这些化合物干扰了ConA处理的人子宫颈成纤维细胞中的前MMP-2激活。尽管如此，弗林蛋白酶反义寡核苷酸完全抑制了兔和人成纤维细胞中的弗林蛋白酶基因表达。这些结果表明弗林蛋白酶不参与ConA诱导的兔真皮成纤维细胞中MT1-MMP的激活过程。
