Cataractogenesis, commonly associated with age, impedes transmission of light to the retina and therefore obstructs vision. Age-related cataracts are managed efficiently by surgical replacement of the aged lens by a man-made lens to restore vision. However, there are no such successful outcomes for the childhood cataracts. Cataracts in infancy/early childhood are debilitating. Their surgical management entails a life-long follow up and impairment of vision. About 50% of the congenital cataracts are genetic and even when there is knowledge about their genetic associations, very little is known about the molecular make-up of the initiating event(s) that lead to the pathogenesis of the cataract. Mutations in the DNA binding domain (DBD) of the heat shock transcription factor, HSF4 have been associated with the most prevalent form of early childhood cataract, the lamellar cataract. Using bacterial artificial chromosome (BAC) transgenesis with manipulated HSF4 (DBD) we have recreated the lamellar cataract pathology in the transgenic mice; thus, for the first time presenting a paradigm for the study of a human pathology that would otherwise be difficult to investigate. In this application we propose to (1) Analyze the transcriptome of the transgenic cataractous lens employing next generation RNA sequencing (RNA-Seq), (2). Analyze the global DNA binding patterns of HSF4 with ChIP-Seq (chromatin immunoprecipitation with next generation DNA sequencing) in the transgenic and the wild type lens and (3). Use the information accrued from aims 1 and 2 to investigate gene expression profiles in individual lens fiber cells by microfluidic qPCR and single cell RNA-sequencing. The approaches of RNA-seq and ChIP-seq and analyses at the single fiber cell level will identify gene(s)/gene expression patterns characteristc of the lamellar cataract pathology. The proposed characterization of individual fiber cells will reveal the nature of the molecular heterogeneity that is associated with fibers, which become cataractous and those that are transparent. But it is not the motivation of this proposal to merely produce a list of genes, the motivation is to contribute to the knowledge base that would lead to (a) exploring pharmaceutical intervention for early childhood cataracts for protecting the visual axis and (b) the establishment of a testable molecular signature that would inform a surgeon's critical decision making to remove or to keep an infant's lens.
Transmission of light to the retina is critical for the neural development in early infancy. A cataract impedes this light transmission. Congenital early childhood cataracts, if left unattended have devastating consequences for the developing child. The ophthalmologist therefore removes the lens as early as possible to protect neural development however; unlike in age-related cataracts the surgical removal of the lens from an infant's eye is beset with lifelong consequences including impairment of vision. With the knowledge of known human mutations we have developed a mouse model directly relevant to the study of the most prevalent form of this childhood pathology, the lamellar cataract. Using this model we will identify genes and molecular pathways associated with the biogenesis of this cataract. This knowledge will lead to the exploration of therapeutic intervention for this debilitating disease and the development of critical decision making information for the surgeon.
| Gangalum, Rajendra K; Kim, Dongjae; Kashyap, Raj K et al. (2018) Spatial Analysis of Single Fiber Cells of the Developing Ocular Lens Reveals Regulated Heterogeneity of Gene Expression. iScience 10:66-79 |
| Gangalum, Rajendra K; Bhat, Ankur M; Kohan, Sirus A et al. (2016) Inhibition of the Expression of the Small Heat Shock Protein ?B-Crystallin Inhibits Exosome Secretion in Human Retinal Pigment Epithelial Cells in Culture. J Biol Chem 291:12930-42 |
