The carboxyl terminal domain of phosducin functions as a transcriptional activator

In previous work, we identified a set of phosducin (Phd) isoforms with unkn own function including the phosducin (Phd) like orphan protein 1 (PhLOP1), an amino terminal truncated isoform of the retinal Phd lacking the G beta g amma binding domain. To investigate the potential biological function of Ph LOP1, PhLOP1 was fused at its amino terminus with the DNA binding domain (B D) of the yeast transcriptional factor, GAL4, and used as bait in a yeast t wo-hybrid screen. Two potential functional protein partners were identified during the screen: SUG1, a subunit of the 265 proteasome and a putative tr anscriptional mediator, and CRX, a retina- and pineal-specific transcriptio n factor. Upon localizing the interacting domain of PhLOP1 with one of the new partners, SUG1, we found that a domain of 40 amino acids at the carboxy l terminus of Phd and PhLOP1 had intrinsic transcriptional activation activ ity in yeast. The transactivation activity was further confirmed in mammali an cells. This region contains an acidic domain that has been shown to be i nvolved in the function of several transcriptional activators. In addition, we showed that Phd is cytoplasmic while PhLOP1 is localized predominantly to the nucleus when fused to an enhanced green fluorescent protein (EGFP) a nd transiently expressed in transfected cells, suggesting that PhLOP1 may p lay a distinct functional role in transcriptional regulation independent of the known Phd interaction/regulation of G beta gamma (C) 2000 Academic Pre ss.