Tissue-specific and high-level expression of the human tyrosine hydroxylase gene in transgenic mice

doi:10.1016/0896-6273(91)90061-4

Neuron

Volume 6, Issue 4, April 1991, Pages 583-594

https://doi.org/10.1016/0896-6273(91)90061-4 Get rights and content

Abstract

Transgenic mice carrying multiple copies of the human tyrosine hydroxylase (TH) gene have been produced. The transgenes were transcribed correctly and expressed specifically in brain and adrenal gland. The level of human TH mRNA in brain was about 50-fold higher than that of endogenous mouse TH mRNA. In situ hybridization demonstrated an enormous region-specific expression of the transgene in substantia nigra and ventral tegmental area. TH immunoreactivity in these regions, though not comparable to the increment of the mRNA, was definitely increased in transgenic mice. This observation was also supported by Western blot analysis and TH activity measurements. However, catecholamine levels in transgenics were not significantly different from those in nontransgenics. These results suggest unknown regulatory mechanisms for human TH gene expression and for the catecholamine levels in transgenic mice.

References (44)

M.M. Bradford
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding
Anal. Biochem.
(1976)
G.T. Coker et al.
Characterization of rat and human tyrosine hydroxylase genes: functional expression of both promoters in neuronal and non-neuronal cell types
Biochem. Biophys. Res. Commun.
(1988)
J.P. Hammang et al.
Immortalized retinal neurons derived from SV40 T-antigen-induced tumors in transgenic mice
Neuron
(1990)
H. Ichinose et al.
Isolation and characterization of a cDNA clone encoding human aromatic l-amino acid decarboxylase
Biochem. Biophys. Res. Commun.
(1989)
N. Kaneda et al.
Isolation of a. novel cDNA clone for human tyrosine hydroxylase: alternative RNA splicing produces four kinds of mRNA from a single gene
Biochem. Biophys. Res. Commun.
(1987)
N. Kaneda et al.
Molecular cloning of cDNA and chromosomal assignment of the gene for human phenylethanolamine N-methyltransferase, the enzyme forepinephrine biosynthesis
J. Biol. Chem.
(1988)
K. Kobayashi et al.
Expression of four types of human tyrosine hydroxylase in COS cells
FEBS Lett.
(1988)
A.R. Krainer et al.
Normal and mutant human β-globin pre-mRNAs are faithfully and efficiently spliced in vivo
Cell
(1984)
E.J. Lewis et al.
Regulation of tyrosine hydroxylase mRNA by glucocorticoid and cyclic AMP in a rat pheochromocytoma cell line
J. Biol. Chem.
(1983)
I.N. Mefford et al.
Simultaneous determination of catecholamine and unconjugated 3,4-dihydroxyphenylacetic acid in brain tissue by ion-pairing reverse-phase high-performance liquid chromatography with electrochemical detection
Anal. Biochem.
(1980)

P.L. Mellon et al.

Immortalization of hypothalamic GnRH neurons by genetically targeted tumorigenesis

Neuron

(1990)

I. Nagatsu et al.

Transient tyrosine hydroxylase-immunoreactive neurons in the region of the anterior olfactory nucleus of pre- and postnatal mice do not contain dopamine

Brain Res.

(1990)

T. Nagatsu et al.

Tyrosine hydroxylase: the initial step in norepinephrine biosynthesis

J. Biol. Chem.

(1964)

T. Nagatsu et al.

Highly sensitive assay for tyrosine hydroxylase activity by high-performance liquid chromatography

J. Chromatogr.

(1979)

K. Oka et al.

Kinetic properties of tyrosine hydroxylase purified from bovine adrenal medulla and bovine caudate nucleus

Biochim. Biophys. Acta

(1982)

T. Sasaoka et al.

Structure of human phenylethanolamine N-methyltransferase gene: existence of two types of mRNA with different transcription initiation sites

Neurochem. Int.

(1989)

A. Berod et al.

Modulation of tyrosine hydroxylase gene expression in the central nervous system visualized by in situ hybridization

F. Cambi et al.

5′ flanking DNA sequences direct cell-specific expression of rat tyrosine hydroxylase

J. Neurochem.

(1989)

K. Chada et al.

Specific expression of a foreign β-globin gene in erythroid cells of transgenic mice

Nature

(1985)

J.M. Chirgwin et al.

Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease

Biochemistry

(1979)

E. Gizang-Ginsberg et al.

Nerve growth factor regulates tyrosine hydroxylase gene transcription through a nucleoprotein complex that contains c-Fos

Genes Dev.

(1990)

J.W. Gordon et al.

Genetic transformation of mouse embryos by microinjection of purified DNA

Cited by (109)

Overexpression of tyrosine hydroxylase in dopaminergic neurons increased sensitivity to methamphetamine
2023, Neurochemistry International
Drug abuse is one of the great social problems in the world and a major healthcare challenge. It is supposed that sensitivity and reactivity to abuse drugs may vary from person to person, while its molecular basis is largely unknown. Dopaminergic neurons are deeply involved in addiction, and tyrosine hydroxylase (TH) catalyzes the first and rate-limiting step of the biosynthesis of dopamine (DA). We investigated the effects of increased TH expression on the metabolism of DA and reactivity to methamphetamine (METH), a drug of abuse, in mice. Wild-type TH (WT-TH) or the S40E mutant of TH (S40E-TH), which is an active form of TH mimicking phosphorylated TH at the 40th serine, was expressed in midbrain dopaminergic neurons using an adeno-associated virus (AAV) vector. The biochemical analysis showed that the turnover rates of DA in the nerve terminals were increased by the expression of WT-TH and S40E-TH, while there were few changes in the DA contents. Next, we administered METH to TH-overexpressing mice. We found that the S40E-TH-expressing mice responded to lower doses of METH than the control mice and WT-TH mice. The stereotyped behaviors appeared first in S40E-TH mice and then in WT-TH and control mice in this order. These data showed that the TH activity and expression level differentially affect DA metabolism in the nerve terminals from that in the cell bodies and that the TH activity and expression level are one of the determining factors for sensitivity and reactivity to METH. We suggest that TH may be a drug target for ameliorating sensitivity to drugs of abuse.
L-DOPA-induced dyskinesia is associated with a deficient numerical downregulation of striatal tyrosine hydroxylase mRNA-expressing neurons
2016, Neuroscience
Citation Excerpt :
A similar regulatory response with steady TH mRNA levels but elevated TH protein expression has been described in individual cell bodies of surviving nigral DAergic neurons in early symptomatic MPTP-treated mice (Kozina et al., 2014). The fact that mechanisms other than transcription may predominantly control general levels of TH and the subsequent biosynthesis of neurotransmitter has already been reported in 1991 by Kaneda et al. (1991) showing TH transgenic mice with a 50-fold increased TH mRNA content but only a two- to threefold elevated TH immunoreactivity and enzyme activity. Interestingly and in contrast to highly active phosphorylated TH in nigral neurons, the enzyme activity of TH in murine intrastriatal neurons has been demonstrated to be low in the state of DAergic denervation, as the TH protein is in a non-serine-phosphorylated state (Depboylu, 2014).
l-3,4-Dihydroxyphenylalanine (l-DOPA) is the therapeutic gold standard in Parkinson’s disease. However, most patients develop debilitating abnormal involuntary movements termed l-DOPA-induced dyskinesia (LID) as therapy-complicating side effects. The underlying mechanisms of LID pathogenesis are still not fully understood. Recent evidence suggests an involvement of striatal tyrosine hydroxylase (TH) protein-expressing neurons, as they are capable of endogenously producing l-DOPA and possibly dopamine. The aim of this study was to elucidate changes of TH transcription in the striatum and nucleus accumbens that occur under experimental conditions of LID.
Mice with a unilateral 6-hydroxydopamine-induced lesion of the medial forebrain bundle were treated daily with l-DOPA for 15 days to provoke dyskinesia. In situ hybridization analysis revealed a significant numerical decrease of TH mRNA-positive neurons in the striatum and nucleus accumbens of mice not exhibiting LID, whereas dyskinetic animals failed to show this reduction of TH transcription. Interestingly, similar changes were observed in intact non-deafferentiated striata, demonstrating an l-DOPA-responsive transcriptional TH regulation independently from nigrostriatal lesion severity.
Consolidation with our previous study on TH protein level (Keber et al., 2015) impressively highlights that LID is associated with both a deficient downregulation of TH transcription and an excessive translation of TH protein in intrastriatal neurons. As TH protein levels in comparison to mRNA levels showed a stronger correlation with development and severity of LID, antidyskinetic treatment strategies should focus on translational and posttranslational modulations of TH as a promising target.
Epigenetic, transcriptional and posttranscriptional regulation of the tyrosine hydroxylase gene
2011, International Journal of Developmental Neuroscience
Citation Excerpt :
The 5 kb of the human TH gene promoter is not sufficient to achieve correct expression of the reporter gene in transgenic mice (Sasaoka et al., 1992). The region located on the 3′ side of the human TH gene exhibits the characteristics of a cell-specific enhancer (Gandelman et al., 1990; Kaneda et al., 1991). The species-specific regulatory site was mapped in the 261-bp region downstream the last exon of the human TH gene.
The activity of tyrosine hydroxylase (TH, EC 1.14.16.2) gene and protein determines the catecholamine level, which, in turn, is crucial for the organism homeostasis. The TH gene expression is regulated by near all possible regulatory mechanisms on epigenetic, transcriptional and posttranscriptional levels. Ongoing molecular characteristic of the TH gene reveals some of the cis and trans elements necessary for its proper expression but most of them especially these responsible for tissue specific expression remain still obscure. This review will focus on some aspects of TH regulation including spatial chromatin organization of the TH locus and TH gene, regulatory elements mediating basal, induced and cell-specific activity, transcriptional elongation, alternative TH RNA processing, and the regulation of TH RNA stability in the cell.
Isolation of an enhancer from the rat tyrosine hydroxylase promoter that supports long-term, neuronal-specific expression from a neurofilament promoter, in a helper virus-free HSV-1 vector system
2007, Brain Research
Citation Excerpt :
However, the binding sites for these transcription factors are located proximal to the transcription start site in the TH promoter, and are not present in the initial upstream fragment of the TH promoter (− 0.6 to − 6.8 kb) that supports long-term expression from the NF-H promoter. Studies in transgenic mice have shown that large fragments of the TH promoter, up to 9 kb, are required for catecholaminergic neuron-specific expression (Banerjee et al., 1992; Kaneda et al., 1991; Min et al., 1994; Sasaoka et al., 1992). Of note, transgenic mice containing 0.15 or 2.4 kb fragments of the rat TH promoter (Min et al., 1994), or 0.2 or 2.5 kb fragments of the human TH promoter (Sasaoka et al., 1992), did not support catecholaminergic neuron-specific expression.
Direct gene transfer into neurons, using a virus vector, has been used to study neuronal physiology and learning, and has potential for supporting gene therapy treatments for specific neurological diseases. Many of these applications require high-level, long-term recombinant gene expression, in forebrain neurons. We previously showed that addition of upstream sequences from the rat tyrosine hydroxylase (TH) promoter to a neurofilament heavy gene (NF-H) promoter supports long-term expression in forebrain neurons, from helper virus-free Herpes Simplex Virus (HSV-1) vectors. This element in the TH promoter satisfied the definition of an enhancer; it displayed activity at a distance from the basal promoter, and in both orientations. This enhancer supported physiological studies that required long-term expression; a modified neurofilament promoter, containing an insulator upstream of the TH-NFH promoter, supported expression in ∼ 11,400 striatal neurons at 6 months after gene transfer, and expression for 7, 8, or 14 months, the longest times tested. In contrast, the NF-H promoter alone does not support long-term expression, indicating that the critical sequences are in the 6.3 kb fragment of the TH promoter. In this study, we performed a deletion analysis to identify the critical sequences in the TH promoter that support long-term expression. We localized these critical sequences to an ∼ 320 bp fragment, and two subfragments of ∼ 100 bp each. Vectors that contained each of these small fragments supported levels of long-term, neuronal-specific expression that were similar to the levels supported by a vector that contained the initial 6.3 kb fragment of the TH promoter. These small fragments of the TH promoter may benefit construction of vectors for physiological studies, and may support studies on the mechanism by which this enhancer supports long-term expression.
Protective effects of neurotrophic factors on tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis of murine adrenal chromaffin cell line tsAM5D
2006, Journal of Biological Chemistry
We previously established the murine adrenal chromaffin cell line tsAM5D, which was immortalized with the temperature-sensitive simian virus 40 large T-antigen. tsAM5D cells have the capacity to differentiate into neuron-like cells in response to neurotrophic factors when the culture temperature is shifted from 33 to 39 °C. In this model system, the temperature shift in the absence of neurotrophic factors led to cell death. Hoechst staining analysis revealed that typical apoptotic nuclei appeared in a time-dependent manner after the temperature shift. Upon shifting to 39 °C, the degradation of T-antigen was accompanied by the transcriptional activation of p53 protein. Among the p53 target genes, death receptor 5 (DR5), which is the receptor for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), showed the highest level of induction. Interestingly, TRAIL-neutralizing antibody protected tsAM5D cells from the temperature shift-induced apoptotic cell death by blocking the activation of caspase-8 and -3, indicating the involvement of TRAIL-mediated death signaling in the temperature shift-induced apoptosis. Glial cell line-derived neurotrophic factor (GDNF) inhibited the TRAIL-mediated activation of caspase-8 in tsAM5D cells exposed to 39 °C and cooperated with basic fibroblast growth factor and ciliary neurotrophic factor. Interestingly, the temperature shift induced oligomerization of DR5, which is the earliest process necessary for transduction of the death signal. This oligomerization was inhibited by treatment with GDNF plus ciliary neurotrophic factor but not by that with GDNF alone or GDNF plus basic fibroblast growth factor. These results are discussed with respect to the intracellular mechanism underlying the protective function of neurotrophic factors against TRAIL-mediated death signaling.
Studies on the Isolation and Molecular Mechanisms of Bioactive Phytochemicals
2022, Yakugaku Zasshi

View all citing articles on Scopus

^†: Present address: Section on Growth Factors, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.

View full text

Neuron

ArticleTissue-specific and high-level expression of the human tyrosine hydroxylase gene in transgenic mice

Abstract

Anal. Biochem.

Biochem. Biophys. Res. Commun.

Neuron

Biochem. Biophys. Res. Commun.

Biochem. Biophys. Res. Commun.

J. Biol. Chem.

FEBS Lett.

Cell

J. Biol. Chem.

Anal. Biochem.

Neuron

Brain Res.

J. Biol. Chem.

J. Chromatogr.

Biochim. Biophys. Acta

Neurochem. Int.

Modulation of tyrosine hydroxylase gene expression in the central nervous system visualized by in situ hybridization

5′ flanking DNA sequences direct cell-specific expression of rat tyrosine hydroxylase

J. Neurochem.

Specific expression of a foreign β-globin gene in erythroid cells of transgenic mice

Nature

Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease

Biochemistry

Nerve growth factor regulates tyrosine hydroxylase gene transcription through a nucleoprotein complex that contains c-Fos

Genes Dev.

Genetic transformation of mouse embryos by microinjection of purified DNA

Article
Tissue-specific and high-level expression of the human tyrosine hydroxylase gene in transgenic mice