TRANSACTIVATION OF THE HUMAN APOLIPOPROTEIN CII PROMOTER BY ORPHAN AND LIGAND-DEPENDENT NUCLEAR RECEPTORS - THE REGULATORY ELEMENT CIIC IS A THYROID-HORMONE RESPONSE ELEMENT

The regulatory elements CIIC (-159/-116) and CIIB (-102-81) of the apo lipoprotein CP (apoCII) promoter have distinct specificities for orpha n nuclear receptors (Vorgia, P., Zannis, V. I., and Kardassis, D. (199 8) J. Biol. Chem. 273, 4188-4199), In this communication we investigat ed the contribution of Ligand-dependent and orphan nuclear receptors o n the transcriptional regulation of the human apoCII gene. It was foun d that element CIIC in addition to ARP-1 and EAR-2 binds RXR alpha/T3R beta heterodimers strongly, whereas element CITE binds hepatic nuclea r factor 4 (HNF-4) exclusively. Binding is abolished by mutations that alter the HRE binding motifs, Transient cotransfection experiments sh owed that in the presence of T3, RXR alpha/T3R beta heterodimers trans activated the -205/+18 apoCII promoter 1.6- and 11-fold in HepG2 and C OS-l respectively. No transactivation was observed in the presence of 9-cis-retinoic acid. Transactivation requires the regulatory element C IIC, suggesting that this element contains a thyroid hormone response element. HNF-4 did not affect the apoCII promoter activity in HepG2 ce lls. However, mutations in the HNF-4 binding site on element CIIB and inhibition of HNF-4 synthesis in HepG2 cells by antisense HNF-4 constr ucts decreased the apoCII promoter activity to 25-40% of the control, indicating that HNF-4 is a positive regulator of the apoCII gene. ARP- 1 repressed the -205/+18 but not the -104/+18 apoCII promoter activity in HepG2 cells, indicating that the repression depends on the regulat ory element CIIC. In contrast, combination of ARP-I and HNF-4 transact ivated different apoCII promoter segments as well as a minimal adenovi rus major late promoter driven by the regulatory element CIIB, Mutagen esis or deletion of elements CIIB or CIIC established that the observe d transactivation requires DNA binding of one of the two factors and m ay result from HNF-4-ARP-1 interactions that elicit the transactivatio n functions of HNF-4. The combined data indicate that RXR alpha/T3R be ta in the presence of T3 and HNF-4 can upregulate the apoCII promoter activity by binding to the regulatory elements CIIC and CIIB, respecti vely. In addition, ARP-1 can either hare inhibitory or stimulatory eff ects on the apoCII promoter activity via different mechanisms.