The regulation of
transcription factors by
kinase or
phosphatase has been well-described. However, little is known about the inactivation of
transcription factors or the nuclear regulators by proteolytic degradation. In this report, we purified a specific
protease,
SPase, from nuclear extracts of the green monkey kidney cell line,
CV-1. Studies of biochemical characteristics and substrate specificity indicated that
SPase is a
cathepsin B-like cysteinyl
protease. However, the two tryptic
peptide sequences derived from the purified
SPase are either identical or highly homologous to those of human
cathepsin L, and furthermore,
SPase shares immunoreactivity with both anti-human
cathepsin L and anti-mouse
cathepsin L antibody. The
SPase was shown to be localized in both cytoplasm and nucleus when subcellular compartments of
CV-1 cells were fractionated.
Transcription factor, SP1, and
retinoblastoma susceptible gene product, RB, are substrates of
SPase while other nuclear factors such as c-Jun and c-Fos are not. These results implied that
SPase plays an integral role in regulating a set of
proteins in the nuclei. In vivo treatment of
CV-1 cells with cysteinyl
protease inhibitor, E-64d, protected RB from degradation.
SPase failed to degrade underphosphorylated RB present in TPA induced terminally differentiated HL-60 or U937 cells. Phosphorylation of RB may cause conformational changes, thus facilitating proteolytic digestion. These observations suggest that an alternative pathway inactivates the function of RB in controlling cell growth. Therefore, a possible role of
SPase may be to affect the stability of important regulators involved in controlling cellular proliferation and differentiation.