Mapping the Universe of Eph Receptor and Ephrin Ligand Transcripts in Epithelial and Fiber Cells of the Eye Lens

Abstract

The eye lens is a transparent, ellipsoid organ in the anterior chamber of the eye that is required for fine focusing of light onto the retina to transmit a clear image. Cataracts, defined as any opacity in the lens, remains the leading cause of blindness in the world. Recent studies in humans and mice indicate that Eph–ephrin bidirectional signaling is important for maintaining lens transparency. Specifically, mutations and polymorphisms in the EphA2 receptor and the ephrin-A5 ligand have been linked to congenital and age-related cataracts. It is unclear what other variants of Ephs and ephrins are expressed in the lens or whether there is preferential expression in epithelial vs. fiber cells. We performed a detailed analysis of Eph receptor and ephrin ligand mRNA transcripts in whole mouse lenses, epithelial cell fractions, and fiber cell fractions using a new RNA isolation method. We compared control samples with EphA2 knockout (KO) and ephrin-A5 KO samples. Our results revealed the presence of transcripts for 12 out of 14 Eph receptors and 8 out of 8 ephrin ligands in various fractions of lens cells. Using specific primer sets, RT-PCR, and sequencing, we verified the variant of each gene that is expressed, and we found two epithelial-cell-specific genes. Surprisingly, we also identified one Eph receptor variant that is expressed in KO lens fibers but is absent from control lens fibers. We also identified one low expression ephrin variant that is only expressed in ephrin-A5 control samples. These results indicate that the lens expresses almost all Ephs and ephrins, and there may be many receptor–ligand pairs that play a role in lens homeostasis.

Spatial-temporal comparison of Eph/Ephrin gene expression in ocular lenses from aging and knockout mice

“…Using traditional, qualitative PCR, 12 of 14 Ephs and 8 of 8 ephrins were previously detected in the lenses of young adult wild-type, EphA2 –/– , and Efna5 –/– mice ( 27 ). Unfortunately, the large number of genes expressed did not sufficiently narrow down targets of interest to pursue.…”

“…Due to the promiscuous nature of Eph-ephrin binding, the number of permutations of receptor-ligand combinations presents a formidable barrier to identify binding partners of interest ( 10 , 25 , 26 ). In the endeavor to narrow targets, a concerted effort to identify the Ephs and ephrins that are present in the mouse lens found that 12 of 14 known Ephs and 8 of 8 known ephrins were expressed ( 27 ). Here, the next step of this search is presented, in which these previously observed Ephs and ephrins are investigated in a quantitative manner.…”

Hotspots and frontiers of genetic research on pediatric cataracts from 2013 to 2022: a scientometric analysis

“…In different species, the disruption of Eph-ephrin signaling was found to be cataractogenesis [23][24][25] . Vu and Cheng [26] extracted and sequenced Eph receptor and ephrin ligand transcripts in adult mice and discovered that practically all Ephs and ephrins were expressed in the adult mouse lens. The spatiotemporal specificity of the expression of other receptor-ligand pairs and their role in the lens, and whether they are affected in the ephrin-A5 mutant lens remain to be elucidated.…”

“…For example, congenital cataracts, a form of cataract in children, are generally characterized by defects in the structure or function of lens proteins and/or their coding genes, which can lead to lens opacity [12] . Utilizing cell biology research methods, we can investigate the pathogenic genes, the abnormalities in the structure of the encoded proteins, and their consequences [15,26] . For instance, changes in isoelectric point or local hydrophobicity/hydrophilicity can cause abnormal aggregation of crystallin, leading to a loss of solubility of high concentrations of intracellular proteins and resulting in lens opacity [12] .…”

Abnormal function of EPHA2/p.R957P mutant in congenital cataract

“…The coordinated elongation and migration of cohorts of lens fiber cells plays an key role in the development of lens sutures [49] . EPHA2-null lenses have displayed the sutural defects and disturbances in the surface patterning of individual fiber cells, implying that EPHA2 has a role in the directed migration of lens fiber cells [50] . Murugan and Cheng [51] reported that the wild-type EPHA2 promoted the migration of mouse lens epithelial αTN4-1 cells in the absence of ligand stimulation, and that the variants in SAM domain diminished this activity.…”