Background: Estrogen acutely activates endothelial nitric oxide synthase (eNOS). However, the identity of the receptors involved in this rapid response remains unclear.
Methods and results: We detected an estrogen receptor alpha (ERalpha) transcript in human endothelial cells that encodes a truncated 46-kDa ERalpha (Delta1a-hERalpha-46). A corresponding 46-kDa ERalpha protein was identified in endothelial cell lysates. Transfection of cDNAs encoding the full-length ERalpha (ERalpha-66) and Delta1a-hERalpha-46 resulted in appropriately sized recombinant proteins identified by anti-ERalpha antibodies. Confocal microscopy revealed that a proportion of both ERalpha-66 and hERalpha-46 was localized outside the nucleus and mediated specific cell-surface binding of estrogen as assessed by FITC-conjugated, BSA-estrogen binding studies. Both ERalpha isoforms colocalized with eNOS and mediated acute activation of eNOS in response to estrogen stimulation. However, estrogen-stimulated transcriptional activation mediated by Delta1a-hERalpha-46 was much less than with ERalpha-66. Furthermore, Delta1a-hERalpha-46 inhibited classical hERalpha-66-mediated transcriptional activation in a dominant-negative fashion.
Conclusions: These findings suggest that expression of an alternatively spliced, truncated ERalpha isoform in human endothelial cells confers a unique ability to mediate acute but not transcriptional responses to estrogen.