ER alpha gene expression in human primary osteoblasts: Evidence for the expression of two receptor proteins

The beneficial influence of E2 in the maintenance of healthy bone is well r ecognized. However, the way in which the actions of this hormone are mediat ed is less clearly understood. Western blot analysis of ER alpha in osteobl asts clearly demonstrated that the well characterized 66-kDa ER alpha was o nly one of the ER alpha isoforms present. Here we describe a 46-kDa isoform of ER alpha, expressed at a level similar to the 66-kDa isoform, that is a lso present in human primary osteoblasts. This shorter isoform is generated by alternative splicing of an ER alpha gene product, which results in exon 1 being skipped with a start codon in exon 2 used to initiate translation of the protein. Consequently, the transactivation domain AF-1 of this ER al pha isoform is absent. Functional analysis revealed that human (h)ER alpha 46 is able to heterodimerize with the full-length ER alpha and also with ER P. Further, a DNA-binding complex that corresponds to hER alpha 46 is detec table in human osteoblasts. We have shown that hER alpha 46 is a strong inh ibitor of hER alpha 66 when they are coexpressed in the human osteosarcoma cell line SaOs. As a functional consequence, proliferation of the transfect ed cells is inhibited when increasing amounts of hER alpha 46 are cotransfe cted with hER alpha 66. In addition to human bone, the expression of the al ternatively spliced ER alpha mRNA variant is also detectable in bone of ER alpha knockout mice. These data suggest that, in osteoblasts, E2 can act in part through an ER a lpha isoform that is markedly different from the 66-kDa receptor. The expre ssion of two ER alpha protein isoforms may account, in part, for the differ ential action that estrogens and estrogen analogs have in different tissues . In particular, the current models of the action of estrogens should be re evaluated to take account of the presence of at least two ER alpha protein isoforms in bone and perhaps in other tissues.