AN INHIBITOR OF NUCLEAR SCAFFOLD PROTEASE BLOCKS CHEMICAL TRANSFORMATION OF FIBROBLASTS

A nuclear scaffold (NS) protease has previously been implicated in pro duction of the M(r) 46,000 ATP-binding protein in NS (which may acquir e nucleoside triphosphatase activity and participate in nucleocytoplas mic transport) by cleavage of a subset of lamins A/C. In a preceding p aper (G. Clawson, L. Norbeck, C. Hatem, C. Rhodes, P. Amiri, J. McKerr ow, S. Patierno, and G. Fiskum, Cell Growth & Differ., 3: 827-838), th is NS protease was identified as a novel, Ca2+-regulated serine protea se, which was found only in the NS and which appears to represent a un ique multicatalytic protease complex. Based upon its predominantly chy motrypsin-like substrate preference, a peptide-chloromethylketone inhi bitor (succinyl-AAPF-chloromethylketone, AAPFcmk) was identified. AAPF cmk showed a K(l) = 56 nm for the NS protease versus 1.4 muM for the e ndoplasmic reticulum activity. Treatment of C3H/10T1/2 mouse embryo fi broblast cells with 1 muM AAPFcmk produced effects which were confined to the nuclear (and to a lesser extent the endoplasmic reticulum) com partment. In this report, we examine the effects of the AAPFcmk inhibi tor on cellular transformation and growth. Growth of C3H/10T1/2 cells was decreased by 34% and 56% at 25 muM and 50 muM AAPFcmk, respectivel y. Growth inhibition occurred without any major change in DNA content distribution, suggesting effects throughout the cell cycle. Growth inh ibition was not observed at lower (less-than-or-equal-to 10 muM) conce ntrations, which decreased transformation of C3H/10T1/2 fibroblasts in a dose-dependent manner by up to 90%, even at femtomolar concentratio ns of AAPFcmk (in the absence of growth inhibition). Inclusion of irre levant inhibitors was without affect. These results suggest that the N S protease may play an important role in acquisition of the transforme d phenotype.