Low-complexity protein domains promote the formation of various biomolecular condensates. However, in many cases, the precise sequence features governing condensate formation and identity remain unclear. Here, we investigate the role of intrinsically disordered mixed-charge domains (MCDs) in nuclear speckle condensation. Proteins composed exclusively of arginine/aspartic-acid dipeptide repeats undergo length-dependent condensation and speckle incorporation.Substituting arginine with lysine in synthetic and natural speckle-associated MCDs abolishes these activities, identifying a key role for multivalent contacts through arginine's guanidinium ion. MCDs can synergise with a speckle-associated RNA recognition motif to promote speckle specificity and residence. MCD behaviour is tuneable through net-charge: increasing negative charge abolishes condensation and speckle incorporation. By contrast, increasing positive charge through arginine leads to enhanced condensation, speckle enlargement, decreased splicing factor mobility, and defective mRNA export. Together, these results identify key sequence determinants of MCD-promoted speckle condensation, and link the speckle's dynamic material properties with function in mRNA processing..
15Low-complexity protein domains promote the formation of various biomolecular 16condensates. However, in many cases, the precise sequence features governing 17 condensate formation and identity remain unclear. Here, we investigate the role of 18 intrinsically disordered mixed-charge domains (MCDs) in nuclear speckle 19 condensation. Proteins composed exclusively of arginine/aspartic-acid dipeptide 20 repeats undergo length-dependent condensation and speckle incorporation. 21Substituting arginine with lysine in synthetic and natural speckle-associated MCDs 22 abolishes these activities, identifying a key role for multivalent contacts through 23 arginine's guanidinium ion. MCDs can synergise with a speckle-associated RNA 24 recognition motif to promote speckle specificity and residence. MCD behaviour is 25 tuneable through net-charge: increasing negative charge abolishes condensation and 26 speckle incorporation. By contrast, increasing positive charge through arginine leads 27 to enhanced condensation, speckle enlargement, decreased splicing factor mobility, 28 and defective mRNA export. Together, these results identify key sequence 29 determinants of MCD-promoted speckle condensation, and link the speckle's dynamic 30 material properties with function in mRNA processing. 31 32 33 34 2015; Nott et al., 2015; Patel et al., 2015). Multivalent interactions among repetitive 58 protein-protein or protein-RNA interaction domains / motifs are the key determinants 59 of LLPS (Dennis, 2015; Fung et al., 2018). Multivalent contacts can be achieved with 60 tandem repeats of folded domains (Li et al., 2012), through low-complexity (LC) 61intrinsically disordered regions (IDRs) (Halfmann, 2016;Lin et al., 2015; Mittag and 62 Parker, 2018), or a combination of the two (Mitrea et al., 2016; Zhang et al., 2015). In 63 these systems, individual contacts are relatively weak. However, with increasing 64 valence, the combined effect of many weak interactions overcome the entropic cost of 65 (Hyman et al., 2014). In many cases LC-IDRs have been shown to be necessary 66 and sufficient for LLPS. This raises key questions regarding the types of residues that 67 promote intermolecular contacts, and the extent to which these determine condensate 68 LLPSResults 103 104 Mixed-charge domains (MCDs) undergo condensation and localize to nuclear 105 speckles 106We previously showed that an arginine and aspartic acid (RD)-enriched MCD 107 from the fungal SPA-5 (SPA-5 MCD ) protein forms a viscoelastic hydrogel. In the fungus, 108 this activity is associated with the formation of cytoplasmic plugs that gate cell-to-cell 109 channels (Lai et al., 2012). To explore the role of such sequences in animal cells, we 110 expressed SPA-5 MCD as an mGFP-fusion in HeLa cells (SPA-5 MCD -mGFP). In this 111 context, SPA-5 MCD -mGFP accumulates in nuclear speckles as revealed by co-112 localization with the speckle marker serine/arginine-rich splicing factor 1 (SRSF1) 113( Figures 1A and S1A). SPA-5 MCD is enriched in RD dipeptide repeats. However, it als...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.