Biology subjects

Jun, A. S.

Publications and source records attributed to Jun, A. S..

3 recordsLinked to original sources

bioRxiv2022

TCF4 trinucleotide repeat expansions and UV irradiation are associated with ferroptosis susceptibility in Fuchs endothelial corneal dystrophy

Fuchs endothelial corneal dystrophy (FECD), the leading indication for corneal transplantation in the U.S., causes loss of corneal endothelial cells (CECs) and corneal edema leading to vision loss. FECD pathogenesis is linked to impaired response to oxidative stress and environmental ultraviolet A (UVA) exposure. Although UVA is known to cause nonapoptotic oxidative cell death resulting from iron-mediated lipid peroxidation, ferroptosis has not been characterized in FECD. We investigated the roles of genetic background and UVA exposure in causing CEC degeneration in FECD. Using ungenotyped FECD patient surgical samples, we found increased levels of cytosolic ferrous iron (Fe2+) and lipid peroxidation in end-stage diseased tissues compared with healthy controls. Using immortalized and primary cell cultures modeling the TCF4 intronic trinucleotide repeat expansion genotype, we found altered gene and protein expression involved in ferroptosis compared to controls including elevated levels of Fe2+, basal lipid peroxidation, and the ferroptosis-specific marker transferrin receptor 1. Increased cytosolic Fe2+ levels were detected after physiologically relevant doses of UVA exposure, indicating a role for ferroptosis in FECD disease progression. Cultured cells were more prone to ferroptosis induced by RSL3 and UVA than controls, indicating ferroptosis susceptibility is increased by both FECD genetic background and UVA. Finally, cell death was preventable after RSL3 induced ferroptosis using solubilized ubiquinol, indicating a role for anti-ferroptosis therapies in FECD. This investigation demonstrates that genetic background and UVA exposure contribute to iron-mediated lipid peroxidation and cell death in FECD, and provides the basis for future investigations of ferroptosis-mediated disease progression in FECD.

cell biology↗

bioRxiv2020

A small molecule that binds an RNA repeat expansion stimulates its decay via the exosome complex

We describe the design of a small molecule that binds the structure of a r(CUG) repeat expansion [r(CUG)exp] and reverses molecular defects in two diseases mediated by the RNA - myotonic dystrophy type 1 (DM1) and Fuchs endothelial corneal dystrophy (FECD). Thus, a single structure-specific ligand has potential therapeutic benefit for multiple diseases, in contrast to oligonucleotide-based modalities that are customized for each disease by nature of targeting the gene that harbors the repeat. Indeed, the small molecule binds the target with nanomolar affinity and >100-fold specificity vs. many other RNAs and DNA. Interestingly, the compounds downstream effects are different in the two diseases, owing to the location of the repeat expansion. In DM1, r(CUG)exp is harbored in the 3 untranslated region (UTR) of and mRNA, and the compound has no effect on the RNAs abundance. In FECD, however, r(CUG)exp is located in an intron, and the small molecule, by binding the repeat expansion, facilitates excision of the intron, which is then degraded by the exosome complex exonuclease, hRRP6. Thus, structure-specific, RNA-targeting small molecules can act disease-specifically to affect biology, either by disabling its gain-of-function mechanism (DM1) or by stimulating quality control pathways to rid a disease-affected cell of a toxic RNA (FECD). Significance statementMany different diseases are caused by toxic structured RNAs. Herein, we designed a lead small molecule that binds a toxic structure and rescues disease biology. We show that a structure-specific small molecule can improve disease-associated defects in two diseases that share the common toxic RNA structure. In one disease, the toxic structure is harbored in an intron and causes its retention. The compound facilitates processing of a retained intron, enabling the disease-affected cell to remove the toxic RNA.

cell biology↗

bioRxiv2020

Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy

A missense mutation of collagen type VIII alpha 2 chain (COL8A2) gene leads to early onset Fuchs endothelial corneal dystrophy (FECD), which progressively impairs vision through loss of corneal endothelial cells. We demonstrate that CRISPR/Cas9-based postnatal gene editing achieves structural and functional rescue in a mouse model of FECD. A single intraocular injection of an adenovirus encoding both the Cas9 gene and guide RNA (Ad-Cas9-Col8a2gRNA), efficiently knocked down mutant COL8A2 expression in corneal endothelial cells, prevented endothelial cell loss, and rescued corneal endothelium pumping function in adult Col8a2 mutant mice. There were no adverse sequelae on histology or electroretinography. Col8a2 start codon disruption represents a non-surgical strategy to prevent vision loss in early-onset FECD. As this demonstrates the ability of Ad-Cas9-gRNA to restore phenotype in adult post-mitotic cells, this method may be widely applicable to adult-onset diseases, even in tissues affected with disorders of non-reproducing cells.

genomics↗