Isoforms of Na,K-ATPase in rats lens epithelium and fiber cells

Purpose. The lens epithelium is thought to conduct Na-K transport for the entire lens cell mass. Lens fibers have a poor ion transport capacity. The authors tested whether different Na,K-ATPase polypeptides are expressed in the two cell types and whether both cells have the machinery needed for ongoing Na,K-ATPase expression as judged by the presence of mRNA for the Na,K-ATPase α subunit. Methods. Membranes were isolated from adult rat lens epithelium or fibers, and Western blot experiments were conducted for Na,K- ATPase α₁, α₂, and α₃ polypeptides. Total RNA was isolated from adult rat lens epithelium or fiber cells, and Northern analysis was conducted for Na,K-ATPase α₁, α₂, and α₃ mRNA. Some experiments were conducted using fiber cells from neonatal (3-day-old) rat lenses. Results. Multiple isoforms of Na,K-ATPase were detected in adult rat lens epithelium. Judged by Northern blot band intensity, mRNA for Na,K-ATPase α₁ and α₂ was more abundant than for α₃ mRNA. By Western blot, Na,K-ATPase α₁, α₂, and α₃ polypeptides were observed as sharp bands at 100 to 108 kDa. In fiber cells, only Na,K-ATPase α₁ immunoreactive polypeptide was detected. Judged by immunoblot density, the amount of α₁ polypeptide was similar in both epithelium and fiber cell material. However, Na,K-ATPase α subunit mRNA was not found in adult lens fibers. To test whether Na,K-ATPase synthesis takes place during fiber cell growth. Northern blot analysis was conducted with mRNA from neonatal (3-day-old) lens fibers: Na,K-ATPase α₁ mRNA was clearly visible. Conclusions. Adult rat lens epithelium expresses more than one isoform of Na,K-ATPase catalytic subunit, whereas only the α₁ isoform can be detected in fiber cells. In adult rat lens fiber cells, the observation of α₁ polypeptide, but no α₁ mRNA, suggests that ongoing α₁ synthesis is low. Based on the detection of α₁ mRNA in neonatal lens fibers, Na,K-ATPase synthesis by lens fibers may be higher during cell elongation and growth.