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Sphingomyelin and RNase-resistant RNA in Microdomains of the Inner Nuclear Membrane

ABSTRACT: Previous evidence has suggested the potential involvement of sphingomyelin and cholesterol located in specific microdomains associated with the inner nuclear membrane in regulating a double-stranded exonuclease-resistant RNA. The objectives of this study were to elucidate the importance of sphingomyelin and cholesterol in safeguarding nuclear RNA from digestion and to scrutinize all RNAs associated with nuclear microdomains. To address these objectives, we investigated the impact of sphingomyelinase or phosphatidylcholine-dependent phospholipase C on nuclear lipid microdomain RNA and subsequently conducted RNA extraction, library preparation, and sequencing of all RNAs present in nuclear microdomains. Only sphingomyelinase treatment rendered the RNA susceptible to RNase treatment. Nuclear lipid microdomains exhibited a higher abundance of retained introns, small nuclear RNA, and long intergenic non-coding RNA compared to whole nuclei, with a notable enrichment in miRNA. The high concentration (20%) of miRNAs in nuclear lipid microdomains was justified by the presence of specific nuclear circular RNA as exons circularized with 'retained' introns, referred to as exon-intron circular RNA (EIciRNA) that act as a sponge of miRNAs. Moreover, it was demonstrated the presence also of ciRNA, circular RNA composed exclusively by introns. The functional analysis indicates that all types of RNase-resistant RNA associated with nuclear lipid microdomains are involved in chromatin organization and brain pathophysiology. In conclusion, nuclear lipid microdomains represent a site of transcription regulation in which circular RNAs, miRNA and double-stranded mRNA, all resistant to RNase, are stabilized by nuclear sphingomyelin.

ORGANISM(S): Mus musculus

PROVIDER: GSE262449 | GEO | 2025/07/24

REPOSITORIES: GEO

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Project description:We have used human umbilical vein endothelial cell line (HUVEC) as a model to investigate the effect of ultrasound (US) activated micro-bubbles on sensitizing the tumour response to radiation and to check the implication of this approach on gene expression. The use of micro-bubbles in cancer therapy is a novel approach that is being recently investigated, and patterns of gene expressions were not yet characterized. Gene analysis has identified about 19,264 genes; where, 239 – 517 genes have shown more than two folds up-regulation in response to the different treatments. This included a number of genes that are known to be involved in apoptosis and ceramide-induced apoptosis pathways, such as Sphingomyelin phosphodiesterase 2, neutral sphingomyelinase (SMPD2), sphingomyelin phosphodiesterase 1, acid lysosomal (ASM; SMPD1), UDP-galactose ceramide galactosyltransferase (UGT8), cytochrome c oxidase (COX6B1), Caspase-9 and mitogen-activated protein kinase kinase 1 (MAP2K1). To verify these results, we have performed Real time RT-PCR, which indicated an up-regulation that ranged from 2 to 200 folds, specifically in the combined treatment of US, 3.3% micro-bubbles and 8 Gy. The observed higher levels of expression of UGT8, SMPD2 and Caspase 9-alpha when compared to the control or to radiation only indicated the involvement of ceramide, which possibly induces apoptosis . These results support the hypothesis that micro-bubbles play a role in increasing sensitization to radiation, and can prove to be an effective cancer therapeutic approach. Gene analysis has identified about 19,264 genes; where, 239 – 517 genes have shown more than two folds up-regulation in response to the different treatments. This included a number of genes that are known to be involved in apoptosis and ceramide-induced apoptosis pathways, such as Sphingomyelin phosphodiesterase 2, neutral sphingomyelinase (SMPD2), sphingomyelin phosphodiesterase 1, acid lysosomal (ASM; SMPD1), UDP-galactose ceramide galactosyltransferase (UGT8), cytochrome c oxidase (COX6B1), Caspase-9 and mitogen-activated protein kinase kinase 1 (MAP2K1). To verify these results, we have performed Real time RT-PCR, which indicated an up-regulation that ranged from 2 to 200 folds, specifically in the combined treatment of US, 3.3% micro-bubbles and 8 Gy. The observed higher levels of expression of UGT8, SMPD2 and Caspase 9-alpha when compared to the control or to radiation only indicated the involvement of ceramide, which possibly induces apoptosis . These results support the hypothesis that micro-bubbles play a role in increasing sensitization to radiation, and can prove to be an effective cancer therapeutic approach.

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Sphingomyelin and RNase-resistant RNA in Microdomains of the Inner Nuclear Membrane

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