Gene · 2025
SRSF10 regulates isoform expression of transcripts associated with proliferative diabetes retinopathy in ARPE-19 cells based on long-read RNA and immunoprecipitation sequencing.
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Jin Siyan, Huang Ju, Wang Yu, Zou He
Аннотация
BACKGROUND: Following injury and disruption of the retinal barrier, retinal pigment epithelium can differentiate into a fibroblastic phenotype, leading to proliferation and migration, thereby resulting in pathological conditions such as proliferative vitreoretinopathy and diabetic retinopathy. Previous studies have detected the specific expression of serine/arginine-rich splicing factor 10 (SRSF10) in the retina; however, its specific function has not been thoroughly studied. SRSF10 has been hypothesized to play an important role in retinal function.
METHODS: We used Oxford Nanopore Technologies (ONT) full-length transcriptome sequencing and Illumina next-generation transcriptome sequencing platforms to detect splice isoforms affected by SRSF10 and employed improved RNA immunoprecipitation sequencing (iRIP-seq) in human retinal pigment epithelial cells to detect RNA binding with SRSF10.
RESULTS: ARPE-19 cells overexpressing SRSF10 showed significant transcriptional differences in the sequencing data obtained from the ONT and Illumina sequencing platforms. Notably, ONT sequencing was more sensitive in detecting new transcripts compared to Illumina. ONT and Illumina sequencing platforms revealed characteristics of alternative splicing events regulated by SRSF10. ONT data showed a primary impact on overlapping exons (olp) events followed by alternative 3' splice site (alt3p) and alt5p, aligning with SRSF10's known functions in exon skipping and inclusion. ONT long-read transcriptome sequencing analysis identified polyadenylation sites (PASs) affected by SRSF10, indicating its role in the dysregulation of polyadenylation in key metabolic genes. In addition, SRSF10 regulates autophagy in cells by influencing the polyadenylation of DEAD-box helicase 58 (DDX58), thereby affecting cell apoptosis.
CONCLUSIONS: The study establishes SRSF10 as a significant splicing factor regulating the alternative splicing of multiple genes and interacting with splicing factors. It plays a pivotal role in cell proliferation, apoptosis, and possibly in the epithelial-mesenchymal transition (EMT) of ARPE-19 cells.
METHODS: We used Oxford Nanopore Technologies (ONT) full-length transcriptome sequencing and Illumina next-generation transcriptome sequencing platforms to detect splice isoforms affected by SRSF10 and employed improved RNA immunoprecipitation sequencing (iRIP-seq) in human retinal pigment epithelial cells to detect RNA binding with SRSF10.
RESULTS: ARPE-19 cells overexpressing SRSF10 showed significant transcriptional differences in the sequencing data obtained from the ONT and Illumina sequencing platforms. Notably, ONT sequencing was more sensitive in detecting new transcripts compared to Illumina. ONT and Illumina sequencing platforms revealed characteristics of alternative splicing events regulated by SRSF10. ONT data showed a primary impact on overlapping exons (olp) events followed by alternative 3' splice site (alt3p) and alt5p, aligning with SRSF10's known functions in exon skipping and inclusion. ONT long-read transcriptome sequencing analysis identified polyadenylation sites (PASs) affected by SRSF10, indicating its role in the dysregulation of polyadenylation in key metabolic genes. In addition, SRSF10 regulates autophagy in cells by influencing the polyadenylation of DEAD-box helicase 58 (DDX58), thereby affecting cell apoptosis.
CONCLUSIONS: The study establishes SRSF10 as a significant splicing factor regulating the alternative splicing of multiple genes and interacting with splicing factors. It plays a pivotal role in cell proliferation, apoptosis, and possibly in the epithelial-mesenchymal transition (EMT) of ARPE-19 cells.