Na+, K+-ATPase inhibitors down-regulate gene expression of the intracellular signaling protein 14-3-3 in rat lens

To identify genes that are differentially expressed by Naf, K+-ATPase inhib itors, we used the differential display technique to compare mRNA expressio n patterns in rat lens. Lenses were treated with 10 mu M ouabain, bufalin, or 19-norbufalin derivative for 24 h and were compared with control lenses. Differential display analysis revealed that one of the downregulated genes was 14-3-3 theta. Down-regulation was confirmed by Northern blot and by re verse transcription-polymerase chain reaction analysis. Reverse transcripti on-polymerase chain reaction of additional 14-3-3 isoforms revealed that th e eta and gamma isoforms of 14-3-3 are also down-regulated by ouabain, bufa lin, and 19-norbufalin derivative, whereas the zeta isoform is downregulate d only by bufalin. Down-regulation of the 14-3-3 isoforms occurred without a significant change in gamma-crystallin gene expression. These results dem onstrate that one of the consequences of Na+, K+-ATPase inhibition by exoge nous or endogenous inhibitors is the down-regulation of mRNA transcripts en coding several isoforms of 14-3-3. Because the 14-3-3 proteins are multifun ctional regulatory proteins, the reduction in the abundance of various isof orms will have profound effects on cell function.