Mapping genome-wide RNA-RNA and RNA-DNA interactions in nuclear hubs of male and female Drosophila cells

Authors

Gunasekera, S., Carlson, M., Ray, M., Larschan, E.

Abstract

Nuclear bodies are nucleoprotein complexes with established functions that target chromatin at specific locations and regulate specific RNA processing functions, thereby influencing gene expression. However, the mechanisms that define how nuclear bodies are targeted to specific locations within the genome where they function remain poorly understood. One significant challenge is capturing and understanding the multiple cell-specific interactions occurring in these complexes, arising from RNA components interacting with each other and with DNA and nucleic acid-binding proteins within the context of the nucleus's three-dimensional organization. Mapping these interactions is critical for elucidating mechanisms such as RNA splicing, a key driver of cell-specific transcript diversity. Here, we use RNA-DNA Split Pool Recognition of Interactions by Tag Extension (RD-SPRITE) to characterize, for the first time, sex-specific RNA-RNA and RNA-DNA interactions in Drosophila S2 (male) and Kc (female) cells. We determined the sex-specific RNA-RNA interaction map within the nucleus and, using RNA-DNA interaction data, pinpointed the target loci of various RNA molecules, including small nuclear RNAs (snRNAs), which are core components of the spliceosome, ribonucleoprotein complex involved in RNA splicing. Based on RNA-RNA interaction data, we also identified novel long non-coding RNAs that may regulate splicing. Furthermore, we investigated the role of transcription factor (TF) CLAMP in sex-specific targeting of the spliceosome. We generated RD-SPRITE datasets in the presence and absence of CLAMP, a key TF involved in dosage compensation, sex-specific RNA splicing, and chromatin organization. We determined that CLAMP regulates global changes in spliceosomal interactions with chromatin, inhibits aberrant snRNA interactions, and regulates sex-specific interactions of RNAs involved in splicing function. Additionally, our dataset provides a valuable resource for investigating additional processes, such as miRNA-mediated silencing, nucleolar functions of snoRNAs, and Cajal body functions of scaRNAs, among others. To facilitate broad community use, we have developed a computational platform, "FlySprite," that enables Drosophila researchers to explore sex-specific RNA-RNA interactions, as well as DNA targets of RNA clusters, through a user-friendly interface.

Preprint server: bioRxiv
The authors list and abstract were imported from bioRxiv on 07 Aug 2026.

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