THE CYCLIC-AMP RESPONSE ELEMENT DIRECTS TYROSINE-HYDROXYLASE EXPRESSION IN CATECHOLAMINERGIC CENTRAL AND PERIPHERAL NERVOUS-SYSTEM CELL-LINES FROM TRANSGENIC MICE

Enhancer elements regulating the neuronal gene, tyrosine hydroxylase ( TH), were identified in TH-expressing peripheral nervous system PATH a nd central nervous system CATH cell lines. Mutational analysis in whic h rat TH 5'-flanking sequences directed chloramphenicol acetyltransfer ase (CAT) reporter gene expression demonstrated that mutating the cycl ic AMP response element (CRE) at -45 base pair reduced expression by 8 0-90%. A CRE linked to an enhancerless TH promoter fully supported exp ression. Cotransfection of a dominant-negative CREB protein reduced ex pression 50-60%, suggesting that the CRE is bound by CREB or a CREB di merization partner. Although mutating the AP1/dyad (AD) element at -20 5 base pair only modestly reduced CAT levels, AD minimal enhancer cons tructs gave 45-80% of wild type expression when positioned at -91 or - 95. However, in its native context at -205, the AD could not support e xpression. In contrast, a CRE, moved from its normal position at -45 t o -206, gave full activity. These results indicate that the CBE is cri tical for TH transcription in central nervous system CATH and peripher al nervous system PATH cells, whereas the AD is less important and its enhancer activity is context- and/or position dependent. These result s represent the first attempts to map regulatory elements directing TH expression in central nervous system cell lines.