Ecdysteroids function through a heterodimeric complex of two nuclear receptors ecdysone receptor (EcR) and ultraspiracle (USP). However the identity of molecules involved in juvenile hormone (JH) action remains elusive. An 85kDa protein identified in Drosophila melanogaster methoprene-tolerant (Met) mutant binds to JH ²²² with high affinity. It is not known whether Met functions by itself or through interactions with other proteins. Reporter assays using yeast LexA two- hybrid system were performed to study the molecular interactions between EcR, USP and Met. EcR fused to LexA DNA binding domain and USP fused to B42 activation domain supported expression of reporter gene even in the absence of ligand. In contrast EcR fused to B42 activation domain and USP fused to LexA DNA binding domain supported lower reporter activity in the absence of ligand and the reporter activity increased upon addition of stable ecdysteroid analog, RG-102240 or steroids muristerone A and ponasterone A. USP:USP homodimers supports expression of reporter gene even in the absence of ligand and there was no significant increase in the reporter activity with the addition of JH analog methoprene. Similarly Met:Met homodimers and Met:EcR and Met:USP heterodimers induced reporter activity in the absence of ligand. Addition of ecdysteroid or JH analogs did not increase the reporter activity regulated by either homodimers or heterodimers of Met protein. These data suggest that the Met protein interacts with EcR and USP in a ligand independent manner.