title: Functional characterization of the S41Y (C2755A) polymorphism of tryptophan hydroxylase 2.
abstract: Human TPH2 (hTPH2) catalyzes the rate-limiting step in CNS serotonin biosynthesis. We characterized a single-nucleotide polymorphism (C2755A) in the hTPH2 gene that substitutes tyrosine for serine at position 41 in the regulatory domain of the enzyme. This polymorphism is associated with bipolar disorder and peripartum depression in a Chinese population. Recombinant h TPH2 human proteins were expressed in bacteria and also stably expressed in PC12 cells. Following bacterial expression and purification, the tyrosine for serine substitution at position 41 (S41Y) polymorphic enzyme displayed increased Vmax with unchanged Km values. By contrast, enzyme stability was decreased in vitro from 32 min to 4 min (37 °C) for the S41Y enzyme (as compared to the wild-type enzyme). The S41Y polymorphism decreased cyclic AMP-dependent protein kinase A-mediated phosphorylation ~ 50% relative to wild-type hTPH2, suggesting that the S41Y mutation may disrupt the post-translational regulation of this enzyme. Transfected PC12 cells expressed hTPH2 mRNA, active protein, and synthesized and released serotonin. Paradoxically, while S41Y-transfected PC12 cells expressed higher levels of hTPH2 than wild type, they synthesized less serotonin. These findings suggest a modified regulation of the S41Y gene variant leading to altered regulation and reduced neurotransmitter synthesis that may contribute to association of the polymorphism with bipolar disorder and depression. We report the functional implications of a polymorphic human tryptophan hydroxylase-2 gene associated with depression and bipolar disorder. The polymorphic enzyme (serine-41 converted to tyrosine) has increased activity, but decreased enzyme stability and serotonin production. Moreover, cyclic AMP-dependent protein kinase (PKA)-mediated phosphorylation of the mutant enzyme is decreased suggesting modified regulation of the S41Y variant leading to altered serotonin.
authors.affiliation: Department of Pharmacology, Penn State College of Medicine, Hershey, Pennsylvania, USA.
authors.forename: Nurgul
authors.initials: N
authors.lastname: Carkaci-Salli
authors.forename: Ugur
authors.initials: U
authors.lastname: Salli
authors.forename: Izel
authors.initials: I
authors.lastname: Tekin
authors.forename: Jeremy A
authors.initials: JA
authors.lastname: Hengst
authors.forename: Moe K
authors.initials: MK
authors.lastname: Zhao
authors.forename: T Lee
authors.initials: TL
authors.lastname: Gilman
authors.forename: Anne M
authors.initials: AM
authors.lastname: Andrews
authors.forename: Kent E
authors.initials: KE
authors.lastname: Vrana
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004298:Dopamine / Q000378:metabolism; D004318:Doxycycline / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D009154:Mutation / Q000235:genetics / Q000502:physiology; D016716:PC12 Cells; D010766:Phosphorylation; D011110:Polymorphism, Genetic / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012701:Serotonin / Q000096:biosynthesis; D013696:Temperature; D014365:Tryptophan Hydroxylase / Q000737:chemistry / Q000235:genetics*
chemical_list: D011994:Recombinant Proteins; D012701:Serotonin; C494784:TPH2 protein, human; D014365:Tryptophan Hydroxylase; D017868:Cyclic AMP-Dependent Protein Kinases; D004318:Doxycycline; D004298:Dopamine
keywords: PC12 cell; TPH2; genetic polymorphism; neurotransmitter biosynthesis; protein stability; serotonin