Cooperative dimerization of the POU domain protein Brn-2 on a new motif activates the neuronal promoter of the human aromatic L-amino acid decarboxylase gene

The neuronal promoter of the human aromatic L-amino acid decarboxylase (AADC) gene contains a perfectly palindromic element (TB) that conforms to the structure of a POU domain protein binding site of the MORE+2 type. The TB motif (located at nts -900/-872 relative to the neuronal cap site) bears striking similarities with the dimeric Pit-1 binding site from growth hormone gene promoter (GH-1), and it enhanced the activity of the minimal tk promoter in transfected SK-N-BE neuroblastoma cells. In transfected COS-7 cells, the expression of a 3xTB-tk-luc was stimulated up to 11-fold by the overexpressed Brn-2 protein. In AADC gene neuronal promoter, we previously characterized a bipartite regulatory element (ONF for octamer-like/NF-Y, nts -86/-57) that binds Brn-2 and NF-Y proteins in a cooperative manner. We now show that both TB and ONF sites participate in the activation of the neuronal promoter by Brn-2. EMSA experiments showed that the recombinant Brn-2 POU domain dimerized on the TB element in a cooperative manner. By site directed mutagenesis of the POU domain of Brn-2, the dimerization interface on the TB element was localized to the hydrophobic pocket of the POU specific domain and the C-terminal part of the POU homeodomain.     

Distribution of AAV2-hAADC-transduced cells after 3 years in Parkinsonian monkeys

The present report describes for the first time, the stability of recombinant adeno-associated virus serotype 2 (AAV2) human aromatic L-amino acid decarboxylase (hAADC) gene transfer after 3-year survival time in a non-human primate model of Parkinson's disease. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned monkeys were treated with six injections of 30 microl/site of AAV2-hAADC at a concentration of 2 x 10(12) vg/ml into the caudate and putamen. Stereological analysis revealed a 46.6% increase in the total number of AAV2-hAADC-transduced cells in the striatum between 8 weeks and 3 years after gene transfer survival time. In the 8-week animals, the distribution of the AADC+ cells was dispersed and heterogeneous, whereas in the 3-year animals it was widespread and homogenous. Confocal analysis demonstrated that approximately 85% of the AADC+ cells were neuronal nuclei immunoreactive.     

Deduced amino acid sequence of bovine aromatic L-amino acid decarboxylase: homology to other decarboxylases

Nucleotide sequence of cDNA for bovine aromatic L-amino acid decarboxylase (AADC) was analyzed. The deduced amino acid sequence of bovine AADC shows 57% identity to drosophila AADC and 37% to plant Catharanthus roseus AADC. The 7-amino acid sequence of the pyridoxal phosphate binding site is completely conserved among drosophila, pig and bovine AADC. AADC primary structure also shows high homology to that of feline glutamic acid decarboxylase.     

Levodopa-responsive aromatic L-amino acid decarboxylase deficiency

We report three siblings, who were treated empirically with levodopa combined with carbidopa. There was an immediate therapeutic response. Biochemical investigation surprisingly showed the clinical phenotype to be caused by aromatic L-amino acid decarboxylase deficiency. Molecular characterization showed a homozygous point mutation (c.387 G-->A) in exon 3. Kinetic studies showed the mutation to decrease the binding affinity for the substrate. This, combined with structural modeling suggesting alteration of active site configuration, provided an explanation for the therapeutic response to levodopa.     

Retinal cholinergic and dopaminergic deficits of aged rats are improved following treatment with GM1 ganglioside

Selected cholinergic and dopaminergic markers were compared in the retina of aged (20–22-months-old) and young (3-months-old) rats before and after treatment with GM1 ganglioside. The dopaminergic markers, tyrosine hydroxylase, aromatic L-amino acid decarboxylase, dopamine, 3,4-dihydroxyphenylacetic acid and homovanillic acid were comparable in the young and aged animals and GM1 treatment did not alter them. In contrast, mazindol binding, a marker for the dopamine transporter, was diminished in the aged retina and treatment with GM1 restored binding to values found in the young animals. The cholinergic markers choline acetyltransferase and hemicholinium-3 binding, a marker for the high-affinity choline transport, were depressed in aged rats and GM1 corrected the deficits.     

Comparative sequencing and association studies of aromatic L-amino acid decarboxylase in schizophrenia and bipolar disorder

Aromatic L-amino acid decarboxylase (AADC) is a relatively non specific enzyme involved in the biosynthesis of several classical neurotransmitters including dopamine and 5-hydroxytryptamine (5HT; serotonin). AADC does not catalyse the rate limiting step in either pathway, but is rate limiting in the synthesis of 2-phenylethylamine (2PE) which is a positive modulator of dopaminergic transmission and a candidate natural psychotogenic compound.1 We and others have proposed that polymorphism in AADC resulting in altered 2PE activity might contribute to the pathogenesis of psychosis. In order to test this hypothesis, we have used denaturing high performance liquid chromatography (DHPLC)3 to screen 3943 bases of the AADC gene and its promoter regions for variants that might affect protein structure or expression in 15 unrelated people with schizophrenia, and 15 unrelated people with bipolar disorder. Three polymorphisms were identified by DHPLC: a insertion/deletion polymorphism in the 5' UTR of the neuronal specific mRNA (g.-33-30delAGAG, bases 586-589 of GenBank M77828), a T>A variant in the non-neuronal exon 1 (g. -67T>A, GenBank M88070), and a G>A polymorphism within intron 8 (g. IVS8 +75G>A, GenBank M84598). Case-control analysis did not suggest that genetic polymorphism in the AADC gene is associated with liability for developing schizophrenia or bipolar disorder.     

Distribution of aromatic L-amino acid decarboxylase mRNA in mouse brain by in situ hybridization histology

Aromatic L-amino acid decarboxylase (AAAD) is the second enzyme in the sequence leading to the synthesis of catecholamines or serotonin. Antisense riboprobes for aromatic L-amino acid decarboxylase mRNA were used to map the gene in mouse brain by in situ hybridization. The substantia nigra, the ventral tegmental nucleus, the dorsal raphe nucleus, the locus coeruleus, and the olfactory bulb contained the highest signal for AAAD mRNA. After treatment with the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), the signal disappeared in the substantia nigra, decreased somewhat in the ventral tegmental area, and remained unchanged in the dorsal raphe nucleus. Hypothalamic and cerebellar Purkinje neurons known to contain histidine decarboxylase or glutamic acid decarboxylase, respectively, were unlabeled by the probes. However, neurons in the deep layers of the frontal cortex, many thalamic nuclei, and the pyramidal neurons of the hippocampus were lightly to moderately labeled for mouse AAAD mRNA. The presence of AAAD message in these neurons suggests that the enzyme has functions other than that for the synthesis of the classical biogenic amine neurotransmitters. © 1993 Wiley-Liss,Inc.