Cloning of a 72 kDa matrix metalloproteinase (gelatinase) from chicken embryo fibroblasts using gene family PCR: expression of the gelatinase increases upon malignant transformation.
Cloning of a 72 kDa matrix metalloproteinase (gelatinase) from chicken embryo fibroblasts using gene family PCR: expression of the gelatinase increases upon malignant transformation.
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使用基因家族 PCR 从鸡胚成纤维细胞中克隆 72 kDa 基质金属蛋白酶（明胶酶）：恶性转化时明胶酶的表达增加。
DOI：
10.1042/bj3000729
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发表时间：
1994
期刊：
影响因子：
--
通讯作者：
Quigley,JP
中科院分区：
文献类型：
--
作者：
Aimes,RT;French,DL;Quigley,JP
鸡胚成纤维细胞分泌一种72 kDa的明胶酶，该酶具有基质金属蛋白酶的所有特征。采用逆转录PCR和简并寡核苷酸引物，是特异性的两个高度保守的序列中发现的所有基质金属蛋白酶，特异性的鸡明胶酶的DNA片段产生。以该PCR产物为探针，从鸡胚cDNA文库中分离cDNA克隆，并测定了整个蛋白质编码序列。在氨基酸水平上，鸡β-胶原酶与人和小鼠的72 kDa β-胶原酶/IV型前胶原酶有84%的相同性，在前肽和催化结构域中存在最大程度的相似性。鸟类和哺乳动物的蛋白酶在C-末端，血红素样结构域有显著差异。鸡明胶酶的最后100个残基与哺乳动物明胶酶只有66%相同。然而，小鼠72 kDa明胶酶在血红素样结构域中与人明胶酶没有显著差异（> 98%同一性）。在这一领域的鸡β-胶原酶的分歧可能解释一些不同的催化和抑制特性的72 kDa的鸡β-胶原酶。Northern-blot分析表明，鸡胚成纤维细胞与劳斯肉瘤病毒（RSV）的恶性转化后，稳定状态水平的鸡β-弹性蛋白酶mRNA增加5倍，3倍，通过治疗与肿瘤促进佛波酯，佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）。这是第一次报道的禽基质金属蛋白酶的克隆。通过RSV转化和肿瘤促进剂PMA增加鸡β-胶原酶的表达表明β-胶原酶在鸡细胞中受到不同的调节。
Chicken embryo fibroblasts secrete a 72 kDa progelatinase that displays all of the characteristics of a matrix metalloproteinase. Employing reverse-transcription PCR and degenerate oligonucleotide primers that are specific for two highly conserved sequences found in all matrix metalloproteinases, a DNA fragment specific for the chicken gelatinase was generated. Using this PCR product as a probe, cDNA clones were isolated from a chicken embryo cDNA library and the entire protein coding sequence was determined. The chicken progelatinase is 84% identical, at the amino acid level, with human and mouse 72 kDa progelatinase/type-IV procollagenase, with the greatest degree of similarity occurring in the propeptide and catalytic domains. The avian and mammalian proteinases diverge significantly in the C-terminal, hemopexin-like domain. The last 100 residues of the chicken gelatinase are only 66% identical with mammalian gelatinases. Mouse 72 kDa progelatinase, however, does not diverge significantly (> 98% identity) from human progelatinase in the hemopexin-like domain. The divergence in this domain of the chicken progelatinase may explain some of the distinct catalytic and inhibitory properties of the 72 kDa chicken progelatinase. Northern-blot analysis reveals that steady-state levels of the chicken progelatinase mRNA are increased 5-fold upon malignant transformation of chicken embryo fibroblasts with Rous sarcoma virus (RSV) and 3-fold by treatment with the tumour-promoting phorbol ester, phorbol 12-myristate 13-acetate (PMA). This represents the first reported cloning of an avian matrix metalloproteinase. The increased expression of the chicken progelatinase by RSV transformation and the tumour promoter PMA suggests that the progelatinase is regulated differently in chicken cells.