IMMUNODETECTION OF MEMBRANE SKELETAL PROTEIN-4.2 IN BOVINE AND CHICKEN EYE LENSES AND ERYTHROCYTES

Purpose. Protein 4.2 is a major erythrocyte membrane skeletal protein, playing an important role in maintaining the integrity and stability of the membrane. It is a transglutaminase-like molecule with no enzyma tic cross-linking activity. Several protein 4.2-associated proteins (i .e. band 3, ankyrin, and protein 4.1) and transglutaminase activities have been detected in the lens. The purpose of this study is to find o ut if protein 4.2 is also expressed in lens fiber membranes. Methods. Western blot analysis of cell membranes isolated from bovine and chick en lens fibers and erythrocytes, and immunocytochemistry of frozen sec tions of bovine and chicken lens fibers were carried out using two pro tein 4.2-specific antibodies. These two peptide antibodies have been u sed to identify two alternatively spliced protein 4.2 isoforms in huma n erythrocyte membranes: the short (P4.2S, or hP4.2(691)) and the long (P4.2L, or hP4.2(721)) isoforms. Results. Western blot analysis using anti-P4.2(L) antibody demonstrated specific immunoreactive polypeptid es in bovine and chicken lens fiber membranes and erythrocyte membrane s, co-migrating with hP4.2(721). Immunofluorescence staining of bovine and chicken lenses, using anti-P4.2(L) antibody, revealed specific si gnals along the cell membranes of cortical fibers. The signals exhibit ed a unique, patchy pattern along the cortical fiber cell membranes in both cross-sectional and longitudinal views. In cross sections, the l abeling of anti-P4.2(L) along the entire cell membranes gave an appear ance of a hexagonal shape of fiber cells. Conclusions. Protein 4.2, or its analogs, is present in the lens fiber membranes. Its specific sta ining pattern in the lens fibers suggests that it participates in the architecture of the lens fiber cell membranes, and may play a role in the lens mechanics and pathology.