IDENTIFICATION OF PHOSPHORYLATION SITES IN THE MOUSE ESTROGEN-RECEPTOR

Phosphorylation sites in the mouse oestrogen receptor, expressed in CO S-1 cells in the presence of 17 beta-oestradiol, have been mapped by s olid phase microsequencing. The receptor was first radiolabelled with [P-32] orthophosphate and a number of H-3- or C-14-labelled amino acid s, immunopurified and then tryptic peptides were separated by thin lay er chromatography or high performance liquid chromatography. Amino aci d sequence analysis indicated that Ser-122, Ser-156, Ser-158 and Ser-2 98 were phosphorylated. The substitution of Ser-122 and Ser-298 with a lanine had a negligible effect on the transcriptional activity of the receptor in transfected cells. However, a reduction of transcriptional activity was observed when Ser-122 was mutated in the context of muta tions in a putative amphipathic a-helix involved in AF-2 activity. Thu s a region of AF-1 that encompasses Ser-122 appears to interact with A F-2 in the full-length receptor.