PT  - JOURNAL ARTICLE
AU  - Jasmine Cubuk
AU  - Jhullian J. Alston
AU  - J. Jeremías Incicco
AU  - Sukrit Singh
AU  - Melissa D. Stuchell-Brereton
AU  - Michael D. Ward
AU  - Maxwell I. Zimmerman
AU  - Neha Vithani
AU  - Daniel Griffith
AU  - Jason A. Wagoner
AU  - Gregory R. Bowman
AU  - Kathleen B. Hall
AU  - Andrea Soranno
AU  - Alex S. Holehouse
TI  - The SARS-CoV-2 nucleocapsid protein is dynamic, disordered, and phase separates with RNA
AID  - 10.1101/2020.06.17.158121
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.06.17.158121
4099  - http://biorxiv.org/content/early/2020/06/18/2020.06.17.158121.short
4100  - http://biorxiv.org/content/early/2020/06/18/2020.06.17.158121.full
AB  - The SARS-CoV-2 nucleocapsid (N) protein is an abundant RNA binding protein that plays a variety of roles in the viral life cycle including replication, transcription, and genome packaging. Despite its critical and multifunctional nature, the molecular details that underlie how N protein mediates these functions are poorly understood. Here we combine single-molecule spectroscopy with all-atom simulations to uncover the molecular details that contribute to the function of SARS-CoV-2 N protein. N protein contains three intrinsically disordered regions and two folded domains. All three disordered regions are highly dynamic and contain regions of transient helicity that appear to act as local binding interfaces for protein-protein or protein-RNA interactions. The two folded domains do not significantly interact with one another, such that full-length N protein is a flexible and multivalent RNA binding protein. As observed for other proteins with similar molecular features, we found that N protein undergoes liquid-liquid phase separation when mixed with RNA. Polymer models predict that the same multivalent interactions that drive phase separation also engender RNA compaction. We propose a simple model in which symmetry breaking through specific binding sites promotes the formation of metastable single-RNA condensate, as opposed to large multi-RNA phase separated droplets. We speculate that RNA compaction to form dynamic single-genome condensates may underlie the early stages of genome packaging. As such, assays that measure how compounds modulate phase separation could provide a convenient tool for identifying drugs that disrupt viral packaging.Competing Interest StatementThe authors have declared no competing interest.