Tissue- and development-specific expression of multiple alternatively spliced transcripts of rat neuronal nitric oxide synthase.

Nitric oxide (NO) functions as an intercellular messenger and mediates numerous biological functions. Among the three isoforms of NO synthase that produce NO, the ubiqui- tously expressed neuronal NO synthase (nNOS) is responsi- ble for a large part of NO production, yet its regulation is poorly understood. Recent reports of two alternative splice- forms of nNOS in the mouse and in man have raised the possibility of spatial and temporal modulation of expres- sion. This study demonstrates the existence of at least three transcripts of the rat nNOS gene designated nNOSa, nNOSb, and nNOSc, respectively, with distinct 5 9 untranslated first exons that arise from alternative splicing to a common sec- ond exon. Expression of the alternative transcripts occurs with a high degree of tissue and developmental specificity, as demonstrated by RNase protection assays on multiple tissues from both fetal and adult rats. Furthermore, termi- nal differentiation of rat pheochromocytoma-derived PC12 cells into neurons is associated with induction of nNOSa, suggesting, likewise, development- and tissue-specific tran- scriptional control of nNOS isoform expression. Physical mapping using a rat yeast artificial chromosome clone shows that the alternatively spliced first exons 1a, 1b, and 1c are separated by at least 15-60 kb from the downstream coding sequence, with exons 1b and 1c being positioned within 200 bp of each other. These findings provide evidence that the biological activ- ity of nNOS is tightly and specifically regulated by a com- plex pattern of alternative splicing, indicating that the no- tion of constitutive expression of this isoform needs to be revised. ( J. Clin. Invest. 1997. 100:1507-1512.) Key words: nitric oxidenitric oxide synthasealternative splicing • gene expressiontranscription