The NPR1 gene of Saccharomyces cerevisiae plays a central role in controlling permease activity; its product is required to promote the activity of at least six distinct transport systems for nitrogenous nutrients under conditions of nitrogen catabolite derepression. We report here the nucleotide sequence of the cloned NPR1 gene. The predicted amino acid sequence indicates that NPR1 encodes a protein of 86 kDa which appears to be organized into two distinct structural domains. The amino-terminal domain of NPR1 (residues 1 to 440) contains 26% serine residues and several regions strongly enriched for PEST residues suggesting a short half-life for the NPR1 protein. The carboxy-terminal region of NPR1 contains consensus sequences characteristic of the catalytic domains of protein kinases. Therefore, NPR1-dependent positive control of nitrogen transport systems most likely involves protein phosphorylation. Northern analysis indicates that the absence of general amino acid permease (GAP1) activity in npr1 mutants is not due to reduction in transcription or messenger stability. Hence, the NPR1 protein probably acts at the post-transcriptional level. Proteins that may serve as substrates for phosphorylation are discussed.