Steroid signaling at the cell surface: Progesterone binding to the N‐terminal extracellular loop of the Rana pipiens oocyte Na/K‐ATPase alpha subunit.

Progesterone (P) activates the amphibian oocyte through a non‐transcriptional signaling system in the plasma membrane that is thought to induce two parallel pathways that converge on the activation of cyclin B‐cdc2. The initial P binding site is a −110 kDa membrane peptide that results in release of a cascade of lipid second messengers and a fall in [cAMP] i . Our analysis of [ 3 H]ouabain binding to the plasma membrane of the Rana pipiens oocyte indicates a high affinity ouabain site linked to pump inhibition and a low affinity ouabain site not associated with the pump. P‐ouabain competition studies indicate that binding of P at the oocyte plasma membrane is at the low affinity ouabain binding site located in the external loop between the H1‐H2 region of the 112 kDa alpha1‐subunit of the Na/K‐ATPase. N and C termini of the alpha‐subunit are located intracellularly and the alpha‐subunit has 10 transmembrane domains. Based on X‐ray crystallographic studies of the P‐cytosol receptor (Nature 393:392, 1998) and site‐directed mutagenesis studies of ouabain binding (Acta Physiol Scand. 163:69, 1998), we predict that the 3‐keto group of P may bind to its peptide receptor via hydrogen bond donors in the external loop between H1‐H2: Gln‐120, pro‐127, Asp‐130 and Asn‐131. Each residue is known to be essential for ouabain inhibition, or identified as determinants of ouabain sensitivity. This may represent the first description of a steroid binding receptor at the cell surface. (Supported by HD 10463).