Altered Cell Clusters and Upregulated Aqp1 in Connexin 50 Knockout Lens Epithemium
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PROVIDER: PRJNA1153492 | ENA |
REPOSITORIES: ENA
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Project description:To characterize the heterogeneity and cell clusters of postnatal lens epithelial cellsLens epithelial cells exhibit an intrinsic heterogeneity of different cell clusters in regulating lens homeostasis and lens growth. Upregulated Aqp1 in Cx50KO lens epithelial cells suggests that both connexin 50 and AQP1 likely play important roles in regulating water homeostasis in lens epithelial cells. (LECs) and to investigate the downstream targets of connexin 50 (Cx50) in the regulation of lens homeostasis and lens growth. To determine differentially expressed genes (DEGs) in the connexin 50 knockout (Cx50KO) lens epithelial cells that shed light on novel mechanism underlying the cataract and small size of the Cx50KO lenses.
2025-09-05 | GSE275886 | GEO
Project description:PurposeTo characterize the heterogeneity and cell clusters of postnatal lens epithelial cells (LECs) and to investigate the downstream targets of connexin 50 (Cx50) in the regulation of lens homeostasis and lens growth. To determine differentially expressed genes (DEGs) in the connexin 50 knockout (Cx50KO) lens epithelial cells that shed light on novel mechanism underlying the cataract and small size of the Cx50KO lenses.MethodsSingle-cell RNA sequencing (scRNA-seq) of lens epithelial cells isolated from one-month-old Cx50KO and wild-type (WT) mice were performed. Differentially expressed genes were identified, and selected DEGs were further studied by quantitative real-time PCR (RT-qPCR) analysis and Western blot analysis.ResultsThe expression profiles of several thousand genes were identified by scRNA-seq data analysis. In comparison to the WT control, many DEGs were identified in the Cx50KO lens epithelial cells, including growth regulating transcriptional factors and genes encoding water channels. Significantly upregulated aquaporin 1 (Aqp1) gene expression was confirmed by RT-qPCR, and upregulated AQP1 protein expression was confirmed by Western blot analysis and immunostaining both in vivo and in vitro.ConclusionsLens epithelial cells exhibit an intrinsic heterogeneity of different cell clusters in regulating lens homeostasis and lens growth. Upregulated Aqp1 in Cx50KO lens epithelial cells suggests that both connexin 50 and AQP1 likely play important roles in regulating water homeostasis in lens epithelial cells.
| S-EPMC11412383 | biostudies-literature
Project description:Connexins 46 and 50 combine to form the gap junctions in ocular lens fiber cells. These proteins are known to be modified with fiber cell age; however, limited work has been done to characterize specific lens connexin modifications. In this report, bovine lens membrane proteins were isolated, digested by multiple enzymes, and analyzed by HPLC-tandem mass spectrometry. Automated database searching revealed the locations of both phosphorylation and truncation sites. The results confirmed the full sequence of connexin 46 and 99% of the connexin 50 sequence. Eighteen phosphorylation sites on connexin 50 and nine phosphorylation sites on connexin 46 were identified, all on serine or threonine residues. All but three phosphorylation sites on connexin 50 were located the cytoplasmic C-terminus. All of the truncation sites of connexin 50 were localized in the cytoplasmic C-terminus (region 280-304). Truncation sites in connexin 46 were found in four different regions including: the N-terminus (residue G2), the cytoplasmic loop (residues 121-124), the cytoplasmic C-terminus (residues 251-285), and the distal C-terminus (residues 344-395). In an analysis of dissected lenses some truncation sites were specific to nucleus samples and others were detected in both nucleus and cortex samples.
| S-EPMC2783236 | biostudies-literature
Project description:Cataracts are mainly classified into three types: cortical cataracts, nuclear cataracts, and posterior subcapsular cataracts. In addition, retrodots and waterclefts are cataract subtypes that cause decreased visual function. To maintain an orderly and tightly packed arrangement to minimize light scattering, adhesion molecules such as connexins and aquaporin 0 (AQP0) are highly expressed in the lens. We hypothesized that some main and/or subcataract type(s) are correlated with adhesion molecule degradation. Lens samples were collected from cataract patients during cataract surgery, and mRNA and protein expression levels were measured by real-time RT-PCR and western blotting, respectively. The mRNA levels of adhesion molecules were not significantly different among any cataract types. Moreover, AQP0 and connexin 46 protein expressions were unchanged among patients. However, connexin 50 protein level was significantly decreased in the lens of patients with WC cataract subtype. P62 and LC3B proteins were detected in the WC patients' lenses, but not in other patients' lenses. These results suggest that more research is needed on the subtypes of cataracts besides the three major types of cataract for tailor-made cataract therapy.
| S-EPMC7718650 | biostudies-literature
Project description:Connexins play essential roles in lens homeostasis and development. Here, we identified a new role for Cx50 that mediates cell-cell adhesion function. Cx50 enhanced the adhesive capability of AQP0. Interestingly, the expression of Cx50 alone promoted cell adhesion at a comparable level to AQP0; however, this cell adhesive function was not observed with other lens connexins, Cx43 and Cx46. Moreover, the adhesive property occurred in both homotypic with Cx50 expressed in both pairing cells and heterotypic with Cx50 in only one pairing cell, and this function appears to be unrelated to its role in forming gap junction channels. Cx50 KO lenses exhibited increased intercellular spaces between lens fiber cells. The second extracellular loop domain (E2) is primarily responsible for this adhesive function. Treatment with a fusion protein containing E2 domain inhibited cell adhesion. Furthermore, disruption of cell adhesion by the E2 domains impaired primary lens cell differentiation. Five critical amino acid residues in the E2 domain primarily are involved in cell adhesive function as well as lens epithelial-fiber differentiation. Together, these results suggest that in addition to forming gap junction channels, Cx50 acts as an adhesive molecule that is critical in maintaining lens fiber integrity and epithelial-fiber differentiation.
| S-EPMC5509658 | biostudies-literature
Project description:Gap junctions establish direct pathways for cell-to-cell communication through the assembly of twelve connexin subunits that form intercellular channels connecting neighbouring cells. Co-assembly of different connexin isoforms produces channels with unique properties and enables communication across cell types. Here we used single-particle cryo-electron microscopy to investigate the structural basis of connexin co-assembly in native lens gap junction channels composed of connexin 46 and connexin 50 (Cx46/50). We provide the first comparative analysis to connexin 26 (Cx26), which-together with computational studies-elucidates key energetic features governing gap junction permselectivity. Cx46/50 adopts an open-state conformation that is distinct from the Cx26 crystal structure, yet it appears to be stabilized by a conserved set of hydrophobic anchoring residues. 'Hot spots' of genetic mutations linked to hereditary cataract formation map to the core structural-functional elements identified in Cx46/50, suggesting explanations for many of the disease-causing effects.
| S-EPMC6309215 | biostudies-literature
Project description:Surgical removal of the lens from larval Xenopus laevis results in a rapid transdifferention of central corneal cells to form a new lens. The trigger for this process is understood to be an induction event arising from the unprecedented contact between the cornea and the vitreous humour that occurs following lens removal. The identity of this trigger is unknown. Here, we have used a functional transgenic approach to show that BMP signalling is required for lens regeneration and a microarray approach to identify genes that are upregulated specifically during this process. Analysis of the array data strongly implicates Wnt signalling and Pitx transcription factors in this process. Pluripotency genes, in contrast, are not upregulated, supporting the idea that corneal cells transdifferentiate without returning to a stem cell state. Furthermore, several genes from the array were expressed in the forming lens during embryogenesis. One of these, nipsnap1, is a known direct target of BMP signalling. We suggest that, as with tail regeneration, activation of multiple developmental signalling pathways could drive lens regeneration from the cornea. Three biological replicates of each of three sample types were analysed. For each replicate, lens tissue (L) was derived from 5 lenses dissected from stage 50 tadpoles. Similarly, 7-8 corneas dissected from stage 50 tadpoles from which the lens had been removed 3 days previously were pooled for each biologocal replicate (R). These samples should contain tissue that is changing from cornea to lens, to regenerate the missing lens. Finally, 7-8 corneas from eyes of tadpoles at the same stage that had the cornea lifetd but the lens left in place 3 days earlier were pooled for each biological replicate (sham operated corneas, designated S).
2011-09-13 | E-GEOD-28014 | biostudies-arrayexpress
Project description: Not available
| EMPIAR-10212 | biostudies-other