The Coronavirus Nucleocapsid Protein Is Dynamically Associated

The coronavirus nucleocapsid (N) protein is a virion structural protein. It also functions, however, in an unknown way in viral replication and localizes to the viral replication-transcription complexes (RTCs). Here we investigated, using recombinant murine coronaviruses expressing green fluorescent protein (GFP)-tagged versions of the N protein, the dynamics of its interactions with the RTCs and the domain(s) involved. Using fluorescent recovery after photobleaching, we showed that the N protein, unlike the nonstructural protein 2, is dynamically associated with the RTCs. Recruitment of the N protein to the RTCs requires the C-terminal N2b domain, which interacts with other N proteins in an RNA-independent manner. All positive-strand RNA viruses assemble their replication complexes in association with intracellular membranes. Coro-naviruses, enveloped plus-strand RNA viruses, induce in in-fected cells the formation of double-membrane vesicles (DMVs) and convoluted membranes (CMs). These structure