Isolation of a collagenase cDNA clone and measurement of changing collagenase mRNA levels during induction in rabbit synovial fibroblasts.

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Abstract
To facilitate studies on the mechanisms controlling collagenase production at a molecular level in rabbit synovial fibroblasts, a c[complementary]DNA library was constructed using mRNA isolated from cells induced with crystals of monosodium urate monohydrate. This library was screened with cDNA probes made from induced and control mRNA populations. From among 30 clones that hybridized preferentially to the induced-cell probe, 4 contained collagenase sequences. The largest, a clone of 650 base pairs, was identified by its ability to hybrid select a mRNA that could be translated in a cell-free system into a product that was precipitable with monospecific antibody to collagenase. Using this clone to probe blots of RNA from induced cells, the appearance of a collagenase mRNA of 2.7 kilobases was detected within 5 h of addition of urate. The level of collagenase mRNA continued to increase for 35-40 h, when it was 60-90 times more abundant in induced cells than in control cells. The increase in mRNA levels correlated with an increase in immunoreactive collagenase protein that was detectable in culture medium by 10 h.