The SAF-A/HNRNPU ATPase and RGG domains are important for XIST RNA localization, nuclear dynamics, RNA splicing, and...

The SAF-A/HNRNPU gene encodes an abundant nuclear protein conserved throughout vertebrates, and is mutated in individuals with HNRNPU syndrome, a neurological human disease. SAF-A is important for maintaining lncRNA localization, splicing, and gene expression state. The mechanistic role of SAF-A in each of these processes is likely coordinated by one or more of its functional domains, which include an N-terminal SAP domain, a central ATPase domain, and an RGG domain defined by a series of C-terminal RGG/RG repeats embedded within a low-complexity region. However, a comprehensive analysis to identify which SAF-A domains are required for each cellular function is lacking. Here, we use an allelic reconstitution strategy to investigate the role of the SAF-A ATPase and RGG domains in lncRNA localization, nuclear dynamics, transcription, splicing, and cell viability. We show that both the ATPase and RGG domains control SAF-A nuclear dynamics, and that SAF-A interacts with nascent RNA Pol II transcripts through the RGG domain. The SAF-A ATPase and RGG domains were required for maintaining XIST RNA and facultative heterochromatin marks on the inactive X chromosome, but did not affect X-linked gene silencing. The SAF-A ATPase and RGG domains were both required for proper mRNA splicing, but not for gene expression. Importantly, we found that the SAF-A ATPase and RGG domains were required for cell proliferation, arguing that these domains are each linked to the essential cellular functions of SAF-A. Collectively, our findings highlight the importance of the SAF-A SAP, ATPase, and RGG domains in vital functions of nuclear biology.