Oxidative stress alters transcript localization of disease-causing genes in the retinal pigment epithelium

Abstract

Nuclear retention is a mechanism whereby RNA transcripts are held in the nucleus to maintain a proper nuclear-to-cytoplasmic balance or as a stockpile for use in responding to stimuli. Many mechanisms are employed to determine whether transcripts are retained or exported to the cytoplasm, though the extent to which tissue- or cell-type, stressors, or disease pathogenesis affect this process remains unclear. As the most biochemically active tissue in the body, the retina must mitigate endogenous and exogenous stressors to maintain cell health and tissue function. Oxidative stress, believed to contribute to the pathogenesis, or progression, of age-related macular degeneration (AMD) and inherited retinal dystrophies (IRDs), is produced both internally from biochemical processes, as well as externally from environmental insult. To evaluate the effect of oxidative stress on transcript localization in the retinal pigment epithelium (RPE), we performed poly-A RNA sequencing on nuclear and cytoplasmic fractions from induced pluripotent stem cell-derived retinal pigment epithelium (iPSC-RPE) cells exposed to hydrogen peroxide, as well as untreated controls. Under normal conditions, the number of mRNA transcripts retained in the nucleus exceeded that found in studies of other tissues. Further, the nuclear-to-cytoplasmic ratio of transcripts is altered following oxidative stress, as is the retention of genes associated with AMD, IRDs, and those important for RPE physiology. These results provide a retention catalog of all expressed mRNA in iPSC-RPE under normal conditions and after exposure to hydrogen peroxide, offering insight into one of the potential roles oxidative stress plays in the progression of visual disorders.

Exploring the lncRNA localization landscape within the retinal pigment epithelium under normal and stress conditions

“…Each sample was sequenced on an Illumina NextSeq 500, and generated a minimum of 13 million reads, with at least 85% of reads uniquely aligned to the hg38 human genome build, and, on average, 92% of these reads counted. We have previously shown these data to be of high quality and the iPSC-RPE to be sufficiently RPE-like [40, 41].…”