Harvard Medical School, New England Primate Research Center, Division of Neurochemistry, One Pine Hill Drive, Southborough, MA 01772-9102, USA.
Tryptophan hydroxylase-2 (TPH2) synthesizes neuronal 5-HT and its genetic variance is associated with numerous behavioral traits and psychiatric disorders. This study characterized the functional significance of two nonsynonymous single nucleotide polymorphisms (SNPs) (C74A and G223A) in rhesus monkey TPH2 (mTPH2). Four haplotypes of mTPH2 were cloned into pcDNA3.1 and stably transfected into PC12 cells. The levels of mTPH2 mRNA and protein were assessed by quantitative real-time PCR and Western blot, respectively, while the intracellular 5-HT was measured by enzyme-linked immunosorbent assay (ELISA). The variant A-A haplotype showed significantly higher levels of mTPH2 mRNA and protein, as well as significantly higher 5-HT production than the wild-type C-G haplotype, while the other two variant haplotypes (C-A and A-G) also tended to produce more 5-HT than C-G haplotype when stably expressed in PC12 cells. Both C74A and G223A were predicted to change mRNA secondary structure, and analysis of the mRNA stability showed that the wild-type C-G haplotype mRNA degrades more quickly than mRNAs of the mutant mTPH2 haplotypes in both stable PC12 and transient HEK-293 cells. This study demonstrates that nonsynonymous SNPs in mTPH2 can affect mRNA stability. Our findings provide an additional mechanism by which nonsynonymous SNPs affect TPH2 function, and further our understanding of TPH2 gene expression regulation.