SPECIFIC-INHIBITION OF THE CHYMOTRYPSIN-LIKE ACTIVITY OF THE PROTEASOME INDUCES A BIPOLAR MORPHOLOGY IN NEUROBLASTOMA-CELLS

Background: Lactacystin inhibits cell proliferation and induces a dist inctive, predominantly bipolar (two-neurite-bearing) morphology in Neu ro 2A murine neuroblastoma cells. It binds with high specificity to th e multicatalytic 20S proteasome and inhibits at least three of its pep tidase activities (chymotrypsinlike, trypsin-like and peptidylglutamyl -peptide hydrolyzing), each at a different rate, without inhibiting ot her known proteases. The chymotrypsin-like and trypsinlike activities of the proteasome are inhibited most rapidly, and irreversibly. In an effort to determine which of the peptidase activities needs to be inhi bited for neurite outgrowth to occur, we treated Neuro 2A cells with p eptide aldehydes that selectively inhibit different proteasome activit ies. Results: Treatment with peptide aldehydes ending in a hydrophobic residue, all of which inhibit the chymotrypsin-like activity, results in a bipolar morphology in Neuro 2A cells, whereas treatment with a p eptide aldehyde inhibitor of the trypsin-like activity does not lead t o a detectable change in morphology. One of the inhibitors that induce s neurite outgrowth has been previously shown to inhibit the chymotryp sin-like activity of the proteasome without inhibiting the other appar ently distinct peptidase activities that cleave after neutral residues , the so-called 'branched chain amino acid preferring' (BrAAP) and 'sm all neutral amino acid preferring' (SNAAP) activities, or the peptidyl glutamyl-peptide hydrolyzing (PGPH) activity. Conclusions: The chymotr ypsin-like activity appears to antagonize bipolar-type neurite outgrow th in Neuro 2A cells, while the trypsin-like, PGPH, BrAAP and SNAAP ap pear not to do so. Selective inhibition of a single peptidase activity , as opposed to general inhibition of the proteasome, appears sufficie nt to induce a specific cellular process. Selective inhibition might b e useful in managing diseases where only one activity is involved with out completely inhibiting the proteasome. It is also possible that end ogenous regulators of the proteasome could affect cellular processes a nd that certain peptidase activities of the proteasome may have roles in specifying a given cell fate.