Modular in vivo assembly of Arabidopsis FCA oligomers into condensates competent for RNA 3' processing.

Our understanding of the functional requirements underpinning biomolecular condensation in vivo is still relatively poor. The Arabidopsis RNA binding protein FLOWERING CONTROL LOCUS A (FCA) is found in liquid-like nuclear condensates that function in transcription termination, promoting proximal polyadenylation at many target genes in the Arabidopsis genome. To further understand the properties of these condensates in vivo, we used single-particle tracking experiments on FCA reporters stably expressed at endogenous levels in plant nuclei. SEC-MALS analyses suggested that FCA forms a core oligomer consistent with a size of four molecules; in vivo particle tracking indicated that this core molecule multimerizes into higher-order particles. The ensuing assemblies coalesce into macromolecular condensates via the coiled-coil protein FLL2, which is genetically required for FCA function. Accordingly, FLL2 predominately co-localizes with FCA in larger-sized condensates. A missense mutation in the FCA RRM domain, also genetically required for FCA function, reduced average size of both FCA particles and condensates, but did not perturb the core oligomer. Our work points to a modular structure for FCA condensates, involving multimerization of core oligomers assembled into functional macromolecular condensates via associated RNA and FLL2 interactions.