Targeted Deletion of the Lens Fiber Cell-Specific Intermediate Filament Protein Filensin

• Published in: Investigative ophthalmology & visual science Vol. 44; no. 12; pp. 5252 - 5258
• Main Authors: Alizadeh, Azita, Clark, John, Seeberger, Teri, Hess, John, Blankenship, Tom, FitzGerald, Paul G
• Format: Journal Article
• Language: English
• Published: Rockville, MD ARVO 01-12-2003; Association for Research in Vision and Ophtalmology
• Subjects: Animals; Biological and medical sciences; Blotting, Northern; Blotting, Western; Cell Differentiation; Chromosome aberrations; Electrophoresis, Polyacrylamide Gel; Eye Proteins - genetics; Eye Proteins - metabolism; Eye Proteins - physiology; Female; Gene Deletion; Gene Targeting; Immunohistochemistry; Intermediate Filament Proteins - genetics; Intermediate Filament Proteins - metabolism; Intermediate Filament Proteins - physiology; Lens diseases; Lens, Crystalline - embryology; Lens, Crystalline - physiology; Lens, Crystalline - ultrastructure; Light; Medical genetics; Medical sciences; Mice; Mice, Inbred C57BL; Mice, Knockout; Ophthalmology; RNA, Messenger - metabolism; Scattering, Radiation; Deletion; Target; Lens; Cell; Protein; Fiber
• ISSN: 0146-0404; 1552-5783; 1552-5783
• DOI: 10.1167/iovs.03-0224

To determine the function of the lens fiber cell-specific cytoskeletal protein, filensin, in lens biology.
Targeted genomic deletion was used to delete exon 1 and the transcriptional start site of the filensin gene. Resultant chimeric animals were bred to homozygosity for the mutant allele. These animals were outbred to mice bearing the wild-type CP49 alleles to eliminate the mutant CP49 gene carried by the 129 strain of mice. Animals homozygous for the mutated filensin gene and wild-type CP49 gene were compared with wild-type and heterozygous animals by Northern and Western blot analyses, light and electron microscopy, and slit lamp microscopy.
Disruption of the filensin gene successfully blocked production of filensin mRNA, reduced levels of filensin's assembly partner CP49, and prevented the assembly of beaded filaments. Despite the absence of beaded filaments, lenses did not show obvious changes in fetal development, nor in the differentiation of epithelial cells into mature fiber cells, as judged by light microscopic analysis. Filensin knockouts began to show evidence of light-scattering by 2 months and worsened with age. Heterozygous animals exhibited an intermediate phenotype, showing a reduction in filensin transcript and moderate light-scattering at 5 months.
The lens fiber cell-specific intermediate filament protein filensin is essential for beaded filament assembly. However, although beaded filaments are not needed for normal lens fetal development or fiber cell differentiation, they appear to be necessary for the long-term maintenance of optical clarity. The mechanism by which the absence of filensin and the beaded filament affects optical clarity has yet to be defined.