A new zebrafish mutant with arrested lens development

Abstract

Abstract: : Purpose: The zebrafish has received considerable attention as a valuable model for genetic studies of vertebrate development. In order to reveal genes essential for lens development, we screened chemically mutagenized zebrafish for related phenotypes and identified a mutation resulting in arrest of lens and anterior segment development, a69. Identification of the gene responsible for a69 may reveal new factors/pathways involved in development of the lens and anterior segment of the eye. Methods: Morphological and histological analysis of zebrafish embryos was performed using standard techniques. Screening of candidate genes was performed by direct sequencing of gene–specific PCR products. Recombinant linkage mapping was performed with simple sequence length polymorphic markers from Research Genetics. Results: The zebrafish a69 mutant is characterized by a malformed eye with a small and often irregular shaped pupil. Another visible defect is a shortened body length. Histological analysis revealed abnormal lens development with onset around 32 hpf. At this stage in normal fish, the developing lens is fully separated from the corneal ectoderm and contains a layer of epithelial cells at the anterior side and differentiating lens fibers. In a69 mutants, the lens cavity is filled with disorganized cells that eventually degenerate. Retinal development also mildly disturbed as photoreceptor markers are found in the inner retina. This is most likely to be a secondary anomaly. Some variability in morphological phenotype was observed depending on genetic background– body length was affected in 98% of ocular mutants in the original "AB" background, while only ∼50% displayed shortened body length in "TU" outcross. Ocular phenotype also varied from mild alterations in the pupil shape and size to severe microphthalmia. Because of the similarity of this phenotype to mouse aphakia and dysgenetic lens, we screened zebrafish pitx3 and foxe3 gene sequences in a69 fish. No mutations have been identified. A genome–wide search for the a69 gene is now underway. Conclusions: From these studies, we conclude that the a69 gene plays an essential role during early stages of lens development and most likely represents a new factor. Identification of the a69 gene will extend our knowledge of factors and mechanisms of vertebrate lens development and facilitate screens for phenotype modifying genes.

Keywords: mutations • anterior segment • gene mapping