The Role of Janus Kinase 3 in the Regulation of Na⁺/K⁺ ATPase under Energy Depletion.

Janus kinase-3 (JAK3) is activated during energy depletion. Energy-consuming pumps include the Na(+)/K(+)-ATPase. The present study explored whether JAK3 regulates Na(+)/K(+)-ATPase in dendritic cells (DCs). Ouabain (100 µM)-sensitive (Iouabain) and K(+)-induced (Ipump) outward currents were determined by utilizing whole cell patch-clamp, Na(+)/K(+)-ATPase α1-subunit mRNA levels by RT-PCR, Na(+)/K(+)-ATPase protein abundance by flow cytometry or immunofluorescence, and cellular ATP by luciferase-assay in DCs from bone marrow of JAK3-knockout (jak3(-/-)) or wild-type mice (jak3(+/+)). Ipump was further determined by voltage clamp in Xenopus oocytes expressing JAK3, active (A568V)JAK3 or inactive (K851A)JAK3. Na(+)/K(+)-ATPase α1-subunit mRNA and protein levels, as well as Ipump and Iouabain were significantly higher in jak3(-/-)DCs than in jak3(+/+)DCs. Energy depletion by 4h pre-treatment with 2,4-dinitro-phenol significantly decreased Ipump in jak3(+/+) DCs but not in jak3(-/-)DCs. Cellular ATP was significantly lower in jak3(-/-)DCs than in jak3(+/+)DCs and decreased in both genotypes by 2,4-dinitro-phenol, an effect significantly more pronounced in jak3(-/-)DCs than in jak3(+/+)DCs and strongly blunted by ouabain in both jak3(+/+) and jak3(-/-)DCs. Ipump and Iouabain in oocytes were decreased by expression of JAK3 and of (A568V)JAK3 but not of (K851A)JAK3. JAK3 inhibitor WHI-P154 (4-[(3'-bromo-4'-hydroxyphenyl)amino]-6,7-dimethoxyquinazoline, 22 μM) enhanced Ipump and Iouabain in JAK3 expressing oocytes. The difference between (A568V)JAK3 and (K851A)JAK3 expressing oocytes was virtually abrogated by actinomycin D (50 nM). JAK3 down-regulates Na(+)/K(+)-ATPase activity, an effect involving gene expression and profoundly curtailing ATP consumption.