PAIRED-LIKE HOMEODOMAIN PROTEINS, PHOX2A AND PHOX2B, ARE RESPONSIBLE FOR NORADRENERGIC CELL-SPECIFIC TRANSCRIPTION OF THE DOPAMINE-BETA-HYDROXYLASE GENE

Recently, a murine paired-like homeobox gene, Phox2a, has been identif ied whose product is critical for the development of several major nor adrenergic neuron populations, including the locus coeruleus. In norad renergic neurons, dopamine P-hydroxylase (DBH) is a hallmark protein a nd catalyzes the conversion of dopamine to noradrenaline, Our previous studies have shown that a composite promoter (domain IV), residing at -185 to -150 bp upstream of the transcription start site, is critical for DBH transcription and is comprised of multiple cis-acting element s, including a cyclic AMP response element, a YY1 binding site, and tw o core motifs of the homeodomain (HD)-binding site. Here, we show that the HD-binding site residing within domain IV is a noradrenergic-spec ific cis-acting element. In contrast, the cyclic AMP response element is active in all cell lines tested. We provide evidence that Phox2a is expressed only in DBH-positive cell lines and interacts with the HD-b inding site. Forced expression of Phox2a robustly activates DBH promot er activity in DBH-negative cell lines (>10-fold), but increased it on ly marginally (<50%) in DBH-positive cell lines. Furthermore, another protein factor with an identical HD, Phox2b, also activates DBH transc ription with an efficiency comparable to that of Phox2a, In contrast, neither Phox2a nor Phox2b was able to transactivate tyrosine hydroxyla se transcription, indicating that these transcription factors differen tially activate catecholamine-synthesizing gene transcription. Togethe r with the Phox2a knockout experiment, the studies described here make Phox2a and Phox2b the first strong candidate transcription factors fo r determining a neurotransmitter phenotype in vertebrates.