A response element for the homeodomain transcription factor Ptx3 in the tyrosine hydroxylase gene promoter

Tyrosine hydroxylase (TH) is the rate-limiting enzyme in the biosynthesis o f catecholamines, which takes place in different types of neuronal systems and nonneuronal tissues. The transcriptional regulation of the TH gene, whi ch is complex and highly variable among different tissues, reflects this he terogeneity. We recently isolated a homeodomain transcription factor, named Ptx3, that is uniquely expressed in the dopaminergic neurons of the substa ntia nigra pars compacta and ventral tegmental area, which together form th e mesencephalic dopaminergic system. This strict localization and its coinc iding induction of expression with the TH gene during development suggested a possible role for this transcription factor in the control of the TH gen e. We report here the presence of a responsive element for Ptx3 located at position -50 to -45 of the rat TH promoter. Transient transfections using T H promoter constructs and electrophoretic mobility shift assays using Ptx3- containing nuclear extracts demonstrated that this region binds Ptx3 protei n and confers a transcriptional effect on the TH gene. Depending on the cel l type, the effect of Ptx3 was an eight- to 12-fold enhancement of TH promo ter activity in Neuro2A neuroblastoma cells, or a 60-80% repression in nonn euronal human embryonic kidney 293 cells. Despite the close association of the Ptx3-binding site and the major cyclic AMP-response element in the TH g ene, no interplay was found between Ptx3 and cyclic AMP-modulating agents. In combination with the orphan nuclear receptor Nurr1, which is required fo r the induction of the TH gene in mesencephalic dopaminergic neurons, the T H promoter activity to Ptx3 was enhanced in Neuro2A cells. Nurr1 alone disp layed only very weak activity on the TH promoter in this cell type. The res ults demonstrate that the homeodomain protein Ptx3 has the potential to act on the promoter of the TH gene in a markedly cell type-dependent fashion. This suggests that Ptx3 contributes to the regulation of TH expression In m esencephalic dopaminergic neurons.