MAMMALIAN PROTEIN RAP46 - AN INTERACTION PARTNER AND MODULATOR OF 70 KDA HEAT-SHOCK PROTEINS

A ubiquitously expressed nuclear receptor-associating protein of simil ar to 46 kDa (RAP46) was identified recently, Interaction experiments with in vitro-translated proteins and proteins contained in cell extra cts revealed that a great variety of cellular regulators associate wit h RAP46, However, in direct interaction tests by the far-Western techn ique, only 70 kDa proteins showed up and were identified as members of the 70 kDa heat shock protein (hsp70) family, Interaction is specific since not all members of the hsp70 family bind to RAP46; interaction occurs through their ATP-binding domain, RAP46 forms complexes with hs p70 in mammalian cells and interacts with hsp70 in the yeast two-hybri d system, Consistent with the fact that hsp70 can bind a multitude of proteins, we identified heteromeric complexes of RAP46-hsp70 with some selected proteins, most notably c-Jun, Complex formation is increased significantly by pre-treatment with alkaline phosphatase, thus sugges ting modulation of interactions by protein phosphorylation. We observe d that RAP46 interferes with efficient refolding of thermally denature d luciferase. Moreover, ATP-dependent binding of misfolded proteins to hsp70 was greatly inhibited by RAP46, These data suggest that RAP46 f unctions as a regulator of hsp70 in higher eukaryotes.