A UNIQUE CATHEPSIN-LIKE PROTEASE ISOLATED FROM CV-1 CELLS IS INVOLVEDIN RAPID DEGRADATION OF RETINOBLASTOMA SUSCEPTIBILITY GENE-PRODUCT, RB, AND TRANSCRIPTION FACTOR SP1

The regulation of transcription factors by kinase or phosphatase has b een well-described. However, little is known about the inactivation of transcription factors or the nuclear regulators by proteolytic degrad ation. In this report, we purified a specific protease, SPase, from nu clear extracts of the green monkey kidney cell line, CV-1. Studies of biochemical characteristics and substrate specificity indicated that S Pase is a cathepsin B-like cysteinyl protease. However, the two trypti c peptide sequences derived from the purified SPase are either identic al or highly homologous to those of human cathepsin L, and furthermore , SPase shares immunoreactivity with both anti human cathepsin L and a nti-mouse cathepsin L antibody. The SPase was shown to be localized in both cytoplasm and nucleus when subcellular compartments of CV-I cell s were fractionated. Transcription factor, SP1, and retinoblastoma sus ceptible gene product, RE, are substrates of SPase while other nuclear factors such as c-Jun and c-Fos are not, These results implied that S Pase plays an integral role in regulating a set of proteins in the nuc lei. In vivo treatment of CV-I cells with cysteinyl protease inhibitor , E-64d, protected RE from degradation. SPase failed to degrade underp hosphorylated RE present in TPA induced terminally differentiated HL-6 0 or U937 cells. Phosphorylation of RE may cause conformational change s, thus facilitating proteolytic digestion. These observations suggest that an alternative pathway inactivates the function of RE in control ling cell growth. Therefore, a possible role of SPase may be to affect the stability of important regulators involved in controlling cellula r proliferation and differentiation.