Cell-specific expression of the human Na+,K(+)-ATPase beta 2 subunit isoform in the nonpigmented ciliary epithelium.
OBJECTIVE
To evaluate the patterns of expression of beta subunit isoforms of the Na+,K(+)-ATPase and H+,K(+)-ATPase in the human eye and to determine the cell-specific distribution of the beta 2 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 (beta 1 and beta 2) or the H+,K(+)-ATPase (alpha and beta) subunits. Antibodies were raised to the amino and carboxyl terminal regions of the human beta 2 isoform. Polymerase chain reaction was used to verify the expression of beta 2 subunit in nonpigmented ciliary epithelial cells (NPE).


RESULTS
Transcripts for the Na+,K(+)-ATPase beta 1 and beta 2 subunit isoforms were present at different levels in all the ocular tissues except the lens, which expressed only beta 1. No transcripts for the alpha or beta subunits of the H+,K(+)-ATPase were detected in the eye. Isoform beta 2 specific anti-peptide antibodies V15E (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 beta 2 isoform is expressed preferentially in the NPE cells. The expression of Na+,K(+)-ATPase beta 2 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 beta 2 subunit isoform, but not H+,K(+)-ATPase, was expressed widely in ocular tissues of the human eye. The restricted cellular distribution of beta 2 isoform within the NPE cells represents an important differential gene marker associated with the multiple alpha subunit isoforms of Na+,K(+)-ATPase.
D000368:Aged; D000369:Aged, 80 and over; D000906:Antibodies / Q000276:immunology; D001483:Base Sequence; D002924:Ciliary Body / Q000201:enzymology*; D003429:Cross Reactions; D004848:Epithelium / Q000201:enzymology; D017506:H(+)-K(+)-Exchanging ATPase / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007223:Infant; D007527:Isoenzymes / Q000378:metabolism*; D008875:Middle Aged; D015335:Molecular Probes; D008969:Molecular Sequence Data; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D013347:Subcellular Fractions / Q000201:enzymology
D000906:Antibodies; D007106:Immune Sera; D007527:Isoenzymes; D015335:Molecular Probes; D017506:H(+)-K(+)-Exchanging ATPase; D000254:Sodium-Potassium-Exchanging ATPase