Cloning and regulation of rat tissue inhibitor of metalloproteinases-2 in osteoblastic cells.
Cloning and regulation of rat tissue inhibitor of metalloproteinases-2 in osteoblastic cells.
复制标题
大鼠组织金属蛋白酶-2抑制剂在成骨细胞中的克隆和调控。
DOI：
10.1006/abbi.1994.1243
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发表时间：
1994
影响因子：
3.9
通讯作者：
Partridge,NC
中科院分区：
文献类型：
--
作者：
Cook,TF;Burke,JS;Bergman,KD;Quinn,CO;Jeffrey,JJ;Partridge,NC
大鼠组织金属蛋白酶抑制剂 2 (TIMP-2) 从 UMR 106-01 大鼠成骨细胞骨肉瘤 cDNA 文库中克隆。 969 bp 全长克隆在氨基酸和核酸水平上与人 TIMP-2 分别具有 98% 和 86% 的序列同一性。暴露 24 小时后，10−8M 的甲状旁腺激素 (PTH) 会刺激 UMR 细胞中 4.2-kb 和 1.0-kb 转录本的基础水平增加约两倍。 TIMP-2 转录物的 PTH 刺激不受蛋白质合成抑制剂放线菌酮 (10−5M) 的影响，表明该激素的主要作用。这与这些相同细胞中 PTH 对间质胶原酶（基质金属蛋白酶-1）的调节形成对比。核连续检测表明，PTH 会导致 TIMP-2 转录增加，与信息水平的增加平行。甲状旁腺激素在刺激 TIMP-2 mRNA 时，似乎通过涉及蛋白激酶 A (PRA) 的信号转导途径发挥作用，因为 TIMP-2 mRNA 的增加是通过 cAMP 类似物 8-bromo-cAMP (5 × 10−3M) 处理再现的。由于佛波醇 l2-肉豆蔻酸酯 13-乙酸酯 (2.6 × 10−6M) 和钙离子载体离子霉素 (10−7M) 对 TIMP-2 转录物丰度缺乏影响，因此蛋白激酶 C 和钙途径似乎不参与其中。在这方面，PTH 对成骨细胞中 TIMP-2 和胶原酶的调节是相似的。然而，我们得出的结论是，由于 TIMP-2 转录的刺激是主要事件，因此 PRA 途径必定负责该基因转录的直接增加。
Rat tissue inhibitor of metalloproteinases-2 (TIMP-2) was cloned from a UMR 106-01 rat osteoblastic osteosarcoma cDNA library. The 969-bp full-length clone demonstrates 98 and 86% sequence identity to human TIMP-2 at the amino acid and nucleic acid levels, respectively. Parathyroid hormone (PTH), at 10−8M, stimulates an approximately twofold increase in both the 4.2-and 1.0-kb transcripts over basal levels in UMR cells after 24 h of exposure. The PTH stimulation of TIMP-2 transcripts was not affected by the inhibitor of protein synthesis, cycloheximide (10−5M), suggesting a primary effect of the hormone. This is in contradistinction to regulation of interstitial collagenase (matrix metalloproteinase-l) by PTH in these same cells. Nuclear run-on assays demonstrate that PTH causes an increase in TIMP-2 transcription that parallels the increase in message levels. Parathyroid hormone, in its stimulation of TIMP-2 mRNA, appears to act through a signal transduction pathway involving protein kinase A (PRA) since the increase in TIMP-2 mRNA is reproduced by treatment with the cAMP analogue, 8-bromo-cAMP (5 × 10−3M). The protein kinase C and calcium pathways do not appear to be involved due to the lack of effect of phorbol l2-myristate 13-acetate (2.6 × 10−6M) and the calcium ionophore, ionomycin (10−7M), on TIMP-2 transcript abundance. In this respect, regulation of TIMP-2 and collagenase in osteoblastic cells by PTH are similar. However, we conclude that since stimulation of TIMP-2 transcription is a primary event, the PRA pathway must be responsible for a direct increase in transcription of this gene.