A common single nucleotide polymorphism alters the synthesis and secretion of neuropeptide Y

A single nucleotide polymorphism (SNP) in the neuropeptide Y gene has been associated with elevated serum lipid levels and cardiovascular disease. The polymorphism (T1128C) changes the seventh amino acid in the prohormone from leucine to proline. It has been speculated this alters neuropeptide Y (NPY) synthesis, trafficking, or secretion. We tested this hypothesis by expressing the mutant and wild-type prohormones in CNS neurons and endocrine cells. Synthesis and trafficking were followed using immunocytochemistry and fluorescent protein-tagged fusion constructs. Mutant prohormone was synthesized and entered the regulated secretory pathway. When expressed in endocrine cells, wild-type and mutant proteins were found in the same large dense core granules. However, the T1128C polymorphism altered the degree of copackaging, and, on average, individual granules contained more mutant prohormone. This was not attributable to codon bias but to the change in prohormone sequence. Global prohormone targeting was normal, because in hippocampal neurons, the polarized distribution of the mutant prohormone was indistinguishable from the wild-type. When secretion was measured from chromaffin cells, brief depolarizations triggered peptide secretion, confirming the entry of the mutant prohormone into the regulated secretory pathway. However, cells that expressed the mutant protein had increased levels of peptide secretion. We conclude that the T1128C polymorphism alters the packaging and secretion of NPY. In contrast to SNPs in other prohormones, we could not find a phenotype until the prohormone was tracked at the single granule level. These results are consistent with studies showing the T1128C polymorphism has pleiotropic effects.