Transcriptional complexity of the Anopheles stephensi nitric oxide synthase gene

Anopheles stephensi nitric oxide synthase (AsNOS) is a single copy gene that shares significant structural homology with the three human NOS genes and is inducibly expressed in Plasmodium-infected mosquitoes. Exon-specific Northern analyses and exon-spanning polymerase chain reaction amplification were used to further characterize transcription from this gene. A total of 18-22 AsNOS transcripts, ranging in size from 1.0 to 7.5 kb, were detected in replicated Northern blots from three separate cohorts of mosquitoes. Three transcripts (1604, 2330, and 2585 bp) were significantly induced in Plasmodium-infected mosquitoes (p<0.05), while others showed varying patterns of induction or downregulation. Five splice variants contained deletions of 1-7 exons. All but one deletion pattern was predicted to introduce in-frame stop codons or alter the translational reading frame. A novel insertion derived from intron sequence was predicted to introduce in-frame stop codons following exon 11. Two truncated novel exon 1 variants were identified that are homologous to a previously published 5' sequence for this exon. The large number of AsNOS transcripts and diversity in AsNOS splicing and exon 1 sequences indicate that transcriptional complexity is a hallmark of both invertebrate and vertebrate NOS genes.