TPH1 (tryptophan hydroxylase 1)

The relationship between occupational stress, musculoskeletal disorders and the mental health of coal miners: The interaction between BDNF gene, TPH2 gene polymorphism and the environment

Tryptophan hydroxylase 2 (TPH2) catalyses the initial and rate‐limiting step in the biosynthesis of serotonin, which is associated with a variety of disorders such as depression, obsessive compulsive disorder, and schizophrenia. Full‐length TPH2 is poorly characterized due to low purification quantities caused by its inherent instability. Three truncated variants of human TPH2 (rch TPH2; regulatory and catalytic domain, NΔ47‐rch TPH2; truncation of 47 residues in the N terminus of rch TPH2, and ch TPH2; catalytic domain) were expressed, purified, and examined for changes in transition temperature, inactivation rate, and oligomeric state. ch TPH2 displayed 14‐ and 11‐fold higher half‐lives compared to rch TPH2 and NΔ47‐rch TPH2, respectively. Differential scanning calorimetry experiments demonstrated that this is caused by premature unfolding of the less stable regulatory domain. By differential scanning fluorimetry, the unfolding transitions of rch TPH2 and NΔ47‐rch TPH2 are found to shift from polyphasic to apparent two‐state by the addition of l‐Trp or l‐Phe. Analytical gel filtration revealed that rch TPH2 and NΔ47‐rch TPH2 reside in a monomer–dimer equilibrium which is significantly shifted toward dimer in the presence of l‐Phe. The dimerizing effect induced by l‐Phe is accompanied by a stabilizing effect, which resulted in a threefold increase in half‐lives of rch TPH2 and NΔ47‐rch TPH2. Addition of l‐Phe to the purification buffer significantly increases the purification yields, which will facilitate characterization of hTPH2.

It was reported recently that male mice lacking brain serotonin (5-HT) lose their preference for females (Liu et al., 2011, Nature, 472, 95–100), suggesting a role for 5-HT signaling in sexual preference. Regulation of sex preference by 5-HT lies outside of the well established roles in this behavior established for the vomeronasal organ (VNO) and the main olfactory epithelium (MOE). Presently, mice with a null mutation in the gene for tryptophan hydroxylase 2 (TPH2), which are depleted of brain 5-HT, were tested for sexual preference. When presented with inanimate (urine scents from male or estrous female) or animate (male or female mouse in estrus) sexual stimuli, TPH2-/- males show a clear preference for female over male stimuli. When a TPH2-/- male is offered the simultaneous choice between an estrous female and a male mouse, no sexual preference is expressed. However, when confounding behaviors that are seen among 3 mice in the same cage are controlled, TPH2-/- mice, like their TPH2+/+ counterparts, express a clear preference for female mice. Female TPH2-/- mice are preferred by males over TPH2+/+ females but this does not lead to increased pregnancy success. In fact, if one or both partners in a mating pair are TPH2-/- in genotype, pregnancy success rates are significantly decreased. Finally, expression of the VNO-specific cation channel TRPC2 and of CNGA2 in the MOE of TPH2-/- mice is normal, consistent with behavioral findings that sexual preference of TPH2-/- males for females is intact. In conclusion, 5-HT signaling in brain does not determine sexual preference in male mice. The use of pharmacological agents that are non-selective for the 5-HT neuronal system and that have serious adverse effects may have contributed historically to the stance that 5-HT regulates sexual behavior, including sex partner preference.

Role of a Serotonin Precursor in Development of Gut Microvilli

A cDNA clone for rabbit tryptophan hydroxylase was used as a probe to identify human tryptophan hydroxylase gene fragments in a panel of hamster-human somatic cell hybrids and determine its chromosomal location in man. A single locus was identified for tryptophan hydroxylase on chromosome 11. Tryptophan hydroxylase is a member of the superfamily of pterin-dependent aromatic amino acid hydroxylases which includes tyrosine hydroxylase, located at 11p15.5-p15, and phenylalanine hydroxylase, located at 12q22-q24.1 in human. The locations of these genes and the evolutionary distance between their sequences suggest that at least three distinct genetic events have occurred during the evolution of the aromatic amino acid hydroxylase superfamily: two sequential gene duplications giving rise to the three distinct hydroxylase loci, and a translocation which separated the tryptophan and tyrosine hydroxylase loci on chromosome 11 from the phenylalanine hydroxylase locus on chromosome 12.

Importance of the field: Serotonin (5-HT) has been implicated in several psychiatric disorders including schizophrenia, depressive disorder and suicide. The key and rate limiting enzyme of 5-HT synthesis is tryptophan hydroxylase 2 (TPH2).Areas covered in this review: The association between TPH2 and affective disorders as well as future vistas of its potential clinical targeting: i) TPH2 in the regulation of 5-HT-dependent behavior, ii) TPH2 gene polymorphism and human behavior, iii) TPH2 and sensitivity to antidepressants and iv) effect of dietary tryptophan manipulation on affective behavior are described.What the reader will gain: The main conclusions of the review are: i) there is an association between TPH2 and genetically defined behavioral variations, ii) the haplotypes, including some human TPH2 gene SNPs, can predict the risk of affective disorders and the sensitivity to antidepressant therapeutics, iii) mutations decreasing TPH2 activity produce negative effects on behavior and, possibly, on survival, iv) the effect of dietary tryptophan manipulations on human mood and behavior is modest compared with that of inhibitors of 5-HT transporter and monoamine oxidase.Take home message: More comprehensive study of TPH2 genetics is needed to increase the clinical value of the enzyme as a predictor of affective disorder risk and efficacy of antidepressant drugs.

Administration of high doses of methamphetamine (METH) produces both short- and long-term enzymatic deficits in central monoaminergic systems. To determine whether a correlative relationship exists between these acute and long-term consequences of METH treatment, in the present study we examined the regional effects of METH on tryptophan hydroxylase (TPH) and tyrosine hydroxylase (TH) activities in various regions of the caudate nucleus, nucleus accumbens, and globus pallidus. A single METH administration decreased TPH activity 1 h after treatment in the globus pallidus, in the nucleus accumbens, and throughout the caudate; in the anterior caudate, the ventral-medial was more affected than the dorsal-lateral region. In contrast, TH activity was not decreased in either the caudate or the globus pallidus after a single METH administration; however, it was altered in the nucleus accumbens. Seven days after multiple METH administrations, TH and TPH activities were decreased in most caudate regions but not in the nucleus accumbens or globus pallidus. These data demonstrate that (1) the effects of METH on TPH and TH vary regionally; and (2) the short-term and long-term regional responses of TPH to METH in the caudate and globus pallidus correlated. In contrast, METH-induced acute TH responses did not predict the long-term changes in TH activity.

Tryptophan hydroxylase 1 (TPH1), which encodes the rate-limiting enzyme tryptophan hydroxylase in the biosynthesis of serotonin, is a candidate gene in the development and treatment response of major depressive disorder (MDD); however, its actual role is uncertain. We aimed to compare the allele frequencies of TPH1 in MDD patients and healthy controls in Taiwan, and also to investigate the association between TPH1 A218C and treatment response to either fluoxetine or venlafaxine in a Taiwanese population with MDD. One hundred five healthy controls and 115 outpatients diagnosed with MDD were recruited and genotyped for the TPH1 218A/C (rs1800532) polymorphism. Patients were randomized into either the fluoxetine or venlafaxine treatment group. The 21-item Hamilton rating scale for depression (HAM-D) was administered to evaluate depressive symptoms at baseline and bi-weekly over 6weeks of treatment. The TPH1 218A/C allele frequencies differed significantly between healthy controls and MDD patients in Taiwan, with a higher prevalence of the A allele in the patient group (p = 0.025). The odds ratio of the A allele to the C allele was 0.507 for the subjects with MDD. There was no significant correlation between the percentage change in HAM-D score and either TPH1 218A/C genotype or TPH1 allele frequencies. This study indicated that the TPH1 218A/C genotype and allele frequencies differed between the Taiwanese healthy controls and MDD patients but could not be used to predict treatment outcome in Taiwanese MDD patients. Further research with larger sample sizes is needed to confirm the role of TPH1 218A/C.

Objective To explore changes of the content of 5-hydrexytryptamine (5-HT) in hippocampns and the relative expression of tryptophan hydroxylase 2 (TPH2) in raphe nuclei in rats of depression model. Methods Twenty female Sprague-Daw]ey rats were randomly divided into two groups (10 in each), normal group and model group, the later was given living alone in combination with 21 day chronic unexpected mild stress (CUMS). Open-field test was used to evaluate behavioral change, the high performance liquid chromatngram-electrocbemical detection, hemi-qnantitative reverse transcriptionpolymerase chain reaction and immunofluorescence were employed to measure 5-HT in hippocampus, to detect the relative expression of TPH2 mRNA and number of TPH2 immunoreactive neurons in raphe nuclei. Results After 21 days of CUMS, beth crossing and rearing in model rats were significantly lower than that in control rats in Open-field test. Compared with the control group, hippocampal 5-HT content was decreased in rats of depression model, and so were the relative expression of TPH2 mRNA and number of TPH2 immunoreactive neurons in raphe nuclei. Conclusion Living alone in combination with 21 days of CUMS could induce rats into depression model, the decrease of hippocampal 5-HT content might result from the decrease of TPH2 expression in raphe nuclei, which led to lower the synthesis of 5-HT. Key words: Stress; Depression; Serotoanin; Tryptophan hydroxylase

The tryptophan hydroxylase 2 (TPH2) enzyme catalyzes the first step of serotonin biosynthesis. Serotonin is known for its role in several homeostatic systems related to sleep, mood, and food intake. As the reaction catalyzed by TPH2 is the rate-limiting step of serotonin biosynthesis, mutations in TPH2 have been associated with several psychiatric disorders (PD). This work undertakes an in silico analysis of the effects of genetic mutations in the human TPH2 protein. Ten algorithms were used to predict the functional and stability effects of the TPH2 mutations. ConSurf was used to estimate the evolutionary conservation of TPH2 amino acids. GROMACS was used to perform molecular dynamics (MD) simulations of TPH2 WT and P260S, R303W, and R441H, which had already been associated with the development of PD. Forty-six TPH2 variants were compiled from the literature. Among the analyzed variants, those occurring at the catalytic domain were shown to be more damaging to protein structure and function. The ConSurf analysis indicated that the mutations affecting the catalytic domain were also more conserved throughout evolution. The variants S364K and S383F were predicted to be deleterious by all the functional algorithms used and occurred at conserved positions, suggesting that they might be deleterious. The MD analyses indicate that the mutations P206S, R303W, and R441H affect TPH2 flexibility and essential mobility at the catalytic and oligomerization domains. The variants P206S, R303W, and R441H also exhibited alterations in dimer binding affinity and stability throughout the simulations. Thus, these mutations may impair TPH2 functional interactions and, consequently, its function, leading to the development of PD. Furthermore, we developed a database, SNPMOL (http://www.snpmol.org/), containing the results presented in this paper. Understanding the effects of TPH2 mutations on protein structure and function may lead to improvements in existing treatments for PD and facilitate the design of further experiments.

Ipsilateral alterations in tryptophan hydroxylase activity in rat brain after hypothalamic 5,7-di-hydroxytryptamine lesion

Tryptophan hydroxylase isoform 2 (TPH2) is expressed in serotonergic neurons in the raphe nuclei, where it catalyzes the rate-limiting step in the synthesis of the neurotransmitter serotonin. In search for functional polymorphisms within the TPH2 gene locus, we measured allele-specific expression of TPH2 mRNA in sections of human pons containing the dorsal and median raphe nuclei. Differences in allelic mRNA expression--referred to as allelic expression imbalance (AEI)--are a measure of cis-acting regulation of gene expression and mRNA processing. Two marker SNPs, located in exons 7 and 9 of TPH2 (rs7305115 and rs4290270, respectively), served for quantitative allelic mRNA measurements in pons RNA samples from 27 individuals heterozygous for one or both SNPs. Significant AEI (ranging from 1.2- to 2.5-fold) was detected in 19 out of the 27 samples, implying the presence of cis-acting polymorphisms that differentially affect TPH2 mRNA levels in pons. For individuals heterozygous for both marker SNPs, the results correlated well (r=0.93), validating the AEI analysis. AEI is tightly associated with the exon 7 marker SNP, in 17 of 18 subjects. Remarkably, expression from the minor allele exceeded that of the major allele in each case, possibly representing a gain-of-function. Genotyping of 20 additional TPH2 SNPs identified a haplotype block of five tightly linked SNPs for which heterozygosity is highly correlated with AEI and overall expression of TPH2 mRNA. These results reveal the presence of a functional cis-acting polymorphism, with high frequency in normal human subjects, resulting in increased TPH2 expression levels. The SNPs that correlate with AEI are closely linked to TPH2 SNPs previously shown to associate with major depression and suicide.

Psychiatric disorders are syndromes characterized by cognitive disturbance and behavioral dysfunction, which affect over 800 million people worldwide. It is considered a major public health problem responsible for severe distress with significant impairment in social and working relationships. In the United States and Canada, psychiatric disorders are considered the main cause of disability in young individuals, in addition to being a key factor underlying suicide. Missense mutations in tryptophan hydroxylase 2 enzyme (TPH2) are associated with the development of psychiatric disorders. TPH2 catalyzes the first step of serotonin biosynthesis, a neurotransmitter that plays a central role in the regulation of emotional behavior and cognition. These mutations lead to TPH2 dysfunction with impaired enzymatic activity, which ultimately results in abnormally low levels of serotonin in the brain. Despite the importance of missense mutations in TPH2 to the development of psychiatric disorders, most of them have not yet been characterized, so their effects are still unknown. In this study, we characterized the impact of missense mutations in TPH2 using prediction algorithms and evolutionary conservation analysis. We also used a penalty system to prioritize the most likely harmful mutations of TPH2 by combining the predictive analyses, evolutionary conservation, literature review, and alterations in physicochemical properties upon amino acid substitution. Three hundred and eighty-four missense mutations of TPH2 were compiled from the literature and databases. Our predictive analysis pointed to a high rate of deleterious and destabilizing predictions for the TPH2 mutations. These mutations mainly affect conserved and, possibly, functionally important residues. Among the uncharacterized mutations of TPH2, variants V295E, R441C T311P, Y281C, R441S, S383F, P308S, Y281H, and E363G received the highest penalties, thus, being the most likely deleterious and, consequently, important targets for future investigation. Our findings may guide the design of clinical and laboratory experiments, optimizing time and resources.

Predominant expression of tryptophan hydroxylase 1 mRNA in the pituitary: A postmortem study in human brain

Alterations in 5-hydroxytryptamine (5-HT) signaling have been implicated as a factor contributing to the altered bowel habit of irritable bowel syndrome (IBS) patients. Tryptophan hydroxylase 1 (TPH1) is the rate-limiting enzyme in enterochromaffin cell 5-HT biosynthesis. We hypothesized that genetic variants affecting TPH1 gene expression might alter intestinal 5-HT bioavailability and subsequently the propensity for distinct bowel habit subtypes in IBS. In this study, we assessed the only common TPH1 proximal promoter variant (-347C/A; rs7130929) and its association with bowel habit predominance in IBS. Electrophoretic mobility shift assays were performed to assess whether the -347C/A-allele variant affects the DNA binding of nuclear factors. Genotype distribution was determined for 422 IBS patients subtyped using the Rome III criteria and for 495 healthy controls recruited from two university medical centers. Association with bowel habit was tested using a multinomial logistic regression model controlling for race, anxiety, depression, and study site. Early growth response factor 1 (EGR-1) bound with higher affinity to a site comprising the minor A-allele of single-nucleotide polymorphism (SNP) -347C/A. TPH1 genotype frequencies did not differ between IBS patients and controls overall. The CC genotype was more prevalent in the IBS-D subtype (47%) than in the IBS-C (25%) and IBS-M (37%) subtypes (P=0.039) after adjusting for race and other covariates. Colonic biopsies from a small cohort of IBS patients from one center were tested for higher TPH1 mRNA expression in samples with CC compared with the CA genotype, but the results did not reach statistical significance. The TPH1 promoter SNP -347C/A differentially binds EGR-1 and correlates with IBS bowel habit subtypes and possibly colonic TPH1 expression consistent with its role in modulating intestinal 5-HT signaling.