Cell-specific expression of the human Na⁺,K⁺-ATPase β2 subunit isoform in the nonpigmented ciliary epithelium

Purpose. To evaluate the patterns of expression of β subunit isoforms of the Na⁺,K⁺-ATPase and H⁺,K⁺-ATPase in the human eye and to determine the cell-specific distribution of the β2 subunit in the human ciliary epithelium. Methods. Total RNA extracted from human ocular tissues was screened by Northern blot analysis with cDNA probes for the human Na⁺,K⁺- ATPase subunit isoforms (β1 and β2) or the H⁺,K⁺-ATPase (α and β) subunits. Antibodies were raised to the amino and carboxyl terminal regions of the human β2 isoform. Polymerase chain reaction was used to verify the expression of β2 subunit in nonpigmented ciliary epithelial cells (NPE). Results. Transcripts for the Na⁺,K⁺-ATPase β1 and β2 subunit isoforms were present at different levels in all the ocular tissues except the lens, which expressed only β1. No transcripts for the α or β subunits of the H⁺,K⁺-ATPase were detected in the eye. Isoform β2 specific anti-peptide antibodies VISE (N-terminus) and A18R (C-terminus) recognized a 55- to 60- kDa protein in the ciliary epithelium and the core protein of 32 kDa after N- glycanase treatment. Immunocytochemical localization within the ciliary epithelium indicates that the Na⁺,K⁺-ATPase β2 isoform is expressed preferentially in the NPE cells. The expression of Na⁺,K⁺-ATPase β2 isoform in the human NPE cell line, ODM-2, was confirmed by polymerase chain reaction amplification and Southern blot analysis. Conclusions. The Na⁺,K⁺- ATPase β2 subunit isoform, but not H⁺,K⁺-ATPase, was expressed widely in ocular tissues of the human eye. The restricted cellular distribution of β2 isoform within the NPE cells represents an important differential gene marker associated with the multiple α subunit isoforms of Na⁺,K⁺-ATPase.