Nuclear speckle-localized RNAs exhibit preferential positioning and orientation

Nuclear speckles are a type of membraneless organelle in higher eukaryotic nuclei. Multivalent interactions between residing proteins and RNAs not only drive the formation, but also the layered compositional organization of nuclear speckles. Here, using super-resolution imaging, we demonstrate that nuclear speckle-localized RNA transcripts exhibit preferential intra-speckle positioning and radial orientation. Specifically, we show that transcripts containing a region enriched in SR protein binding motifs and another region enriched in hnRNP binding motifs localize to the outer shell. Moreover, they are oriented so that the SR enriched region is closer to the speckle center relative to the hnRNP enriched region. This differential intra-speckle positioning of RNA transcripts is driven by multivalent interactions between the two motifs and SR/hnRNP proteins respectively, two protein families that exhibit distinct subcellular localization relative to nuclear speckles. Such intra-speckle RNA positioning and orientation might explain the importance of RNAs in maintaining the structural integrity of nuclear speckles, and point at their possible functional roles in RNA splicing. ### Competing Interest Statement The authors have declared no competing interest.