2 MURINE HOMOLOGS OF THE DROSOPHILA SINGLE-MINDED PROTEIN THAT INTERACT WITH THE MOUSE ARYL-HYDROCARBON RECEPTOR NUCLEAR TRANSLOCATOR PROTEIN

Drosophila single-minded, which acts as a positive master gene regulat or in central nervous system midline formation in Drosophila, its two mouse homologs SIM1 and SIM2, and the mammalian aryl hydrocarbon recep tor (AHR) and aryl hydrocarbon receptor nuclear translocator (ARNT) pr oteins are members of the basic-helix-loop-helix PAS family of transcr iption factors. In the yeast two-hybrid system, we demonstrate strong constitutive interaction of ARNT with SIM1 and SIM2 and fully ligand-d ependent interaction of ARNT with AHR. Both the helix-loop-helix and t he PAS regions of SIM1 and of ARNT are required for efficient heterodi merization. SIM1 and SIM2 do not form homodimers, and they do not inte ract with AHB We also failed to detect homodimerization of ARNT. The i nteraction of ARNT with SIM1 was confirmed with in vitro synthesized p roteins. Like AHR, in vitro synthesized SIM1 associates with the 90-kD a heat shock protein. SIM1 inhibits binding of the AHR . ARNT dimer to the xenobiotic response element in vitro. Introduction of SIM1 into h epatoma cells inhibits transcriptional transactivation by the endogeno us AHR . ARNT dimer. The mouse SIM1 . ARNT dimer binds only weakly to a proposed DNA target for the Drosophila SIM . ARNT dimer. In adult mi ce mRNA for SIM1 was expressed in lung skeletal muscle, and kidney, wh ereas the mRNA for SIM2 was found in the latter two. ARNT is also expr essed in these organs. Thus mouse SIM1 and SIM2 are novel heterodimeri zation partners for ARNT in vitro, and they may function both as posit ive and negative transcriptional regulators in vivo, during embryogene sis and in the adult organism.