Substrate-dependent regulation of human arylamine N-acetyltransferase-1 incultured cells

Arylamine N-acetyltransferase-1 (NAT1) is a polymorphically expressed enzym e that is widely distributed throughout the body. In the present study, we provide evidence for substrate-dependent regulation of this enzyme. Human p eripheral blood mononuclear cells cultured in medium supplemented with p-am inobenzoic acid (PABA; 6 mu M) for 24 h showed a significant decrease (50-8 0%) in NAT1 activity. The loss of activity was concentration-dependent (EC5 0 similar to 2 mu M) and selective because PABA had no effect on the activi ty of constitutively expressed lactate dehydrogenase or aspartate aminotran sferase. PABA also induced down-regulation of NAT1 activity in several huma n cell lines grown at confluence. Substrate-dependent downregulation was no t restricted to PABA. Addition of other NAT1 substrates, such as p-aminosal icylic acid, ethyl-p-aminobenzoate, or p-aminophenol to peripheral blood mo nonuclear cells in culture also resulted in significant (P < .05) decreases in NAT1 activity. However, addition of the NAT2-selective substrates sulfa methazine, dapsone, or procainamide did not alter NAT1 activity. Western bl ot analysis using a NAT1-specific antibody showed that the loss of NAT1 act ivity was associated with a parallel reduction in the amount of NAT1 protei n (r(2) = 0.95). Arylamines that did not decrease NAT1 activity did not alt er NAT1 protein levels. Semiquantitative reverse transcriptase polymerase c hain reaction of mRNA isolated from treated and untreated cells revealed no effect of PABA on NAT1 mRNA levels. We conclude that NAT1 can be down-regu lated by arylamines that are themselves NAT1 substrates. Because NAT1 is in volved in the detoxification/activation of various drugs and carcinogens, s ubstrate-dependent regulation may have important consequences with regard t o drug toxicity and cancer risk.