Arginine-Enriched Mixed-Charge Domains Provide Cohesion for Nuclear Speckle Condensation

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 synergize with a speckle-associated RNA recognition motif to promote speckle specificity and residence. MCD behavior is tunable through net-charge: increasing negative charge abolishes condensation and speckle incorporation. Contrastingly, 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 dynamic material properties of speckles with function in mRNA processing. [Display omitted] •Arginine-enriched mixed-charge domains form condensates and drive speckle assembly•Substituting arginine with lysine abolishes mixed-charge domain (MCD) activity•MCDs synergize with RNA-binding domains to promote speckle residence•Increasing speckle cohesion through MCD expression leads to defects in mRNA export Greig et al. analyze natural and synthetic low-complexity mixed-charge domains (MCDs) to identify key sequence features influencing nuclear speckle condensation and function. Increasing negative charge abolishes MCD activity, whereas increasing positive charge through arginine, but not lysine, leads to increased speckle cohesion, enlarged speckles, and defects in mRNA export.