The nucleus of higher eukaryotes is a highly compartmentalized and dynamic organelle consisting of several biomolecular condensates that regulate gene expression at multiple levels (Banani et al., 2017; Shin and Brangwynne, 2017). First reported more than 100 years ago by Ramón y Cajal, nuclear speckles (NS) are among the most prominent of such condensates (Spector and Lamond, 2011). Despite their prevalence, research on the function of NS is virtually restricted to colocalization analyses, since an organizing core, without which NS cannot form, remains unidentified (Chen and Belmont, 2019; Galganski et al., 2017). The monoclonal antibody SC35, which was raised against a spliceosomal extract, is a frequently used reagent to mark NS since its debut in 1990 (Fu and Maniatis, 1990). Unexpectedly, we found that this antibody has been misidentified and the main target of SC35 mAb is SRRM2, a large (~300 kDa), spliceosome-associated (Jia and Sun, 2018) protein with prominent intrinsically disordered regions (IDRs) that sharply localizes to NS (Blencowe et al., 1994). Here we show that, the core of NS is likely formed by SON and SRRM2, since depletion of SON leads only to a partial disassembly of NS as reported previously (Ahn et al., 2011; Fei et al., 2017; Sharma et al., 2010), in contrast, combined depletion of SON together with SRRM2, but not other NS associated factors, or depletion of SON in a cell line where IDRs of SRRM2 are genetically deleted, leads to a near-complete dissolution of NS. This work, therefore, paves the way to study the role of NS under diverse physiological and stress conditions.
All data generated or analysed during this study are included in the manuscript and Supplementary files and source data files.Mass Spectrometry results shown in Figure 1 and Figure1- figure supplement 1 are provided in Supplementary File 2.Furthermore, on ProteomeXchange with identifier PXD021814.Source data files have been provided for Figure 4, Figure 4 - figure supplement1 and 2; and Figures 5, Figure 5 - figure supplement 3 and 4 .These zipped files contain ilastik models, CellProfiler pipelines, results and Jupyter notebooks.
Orthology and disorder prediction of metazoan SRRM2, SRRM1, SON, Coilin, Pinin, PRPF8 and RBM2510.5281/zenodo.4065244.
SON and SRRM2 form nuclear speckles in human cellsProteomeXchange, PXD021814.
- Tuğçe Aktaş
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Jonathan P Staley, University of Chicago, United States
© 2020, Ilik et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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