The protein encoded by a randomly selected rat brain cDNA clone was identified as neuron-specific enolase (NSE; 126.96.36.199; gamma subunit), based on homology to yeast enolase sequences and the presence of the corresponding 2.5-kb mRNA in rat brain but not in liver, kidney, or muscle tissue. The 2,222-nucleotide NSE and mRNA sequence presented identifies a 68-nucleotide 5' noncoding region, a 1,302-nucleotide open reading frame (corresponding to a primary translation product of 434 amino acids), and 852 noncoding 3' bases. Evolutionary implications based on sequence comparisons to yeast enolase and non-neuronal enolase are discussed. Primer extension analysis indicated the presence of several alternative initiation sites for transcription within 60 nucleotides on the NSE gene. The developmental onset of NSE mRNA expression correlates with the appearance of NSE protein; however, the mRNA reaches adult levels by postnatal week 3, whereas the protein continues to accumulate over the next few months, suggesting regulatory mechanisms in addition to transcriptional control.