What is the Molecular Grammar that Defines Viral Biomolecular Condensates?

Based on similarities to components of cellular biomolecular condensates or membraneless organelles, at least two types of viral protein, namely antiterminators and nucleocapsid proteins, have features and roles consistent with their ability to phase separate with viral RNA molecules.

Antiterminator proteins are a major constituent of viral replication organelles and are often used as markers to visualize these replication compartments in infected cells. Antiterminator proteins typically interact with the viral RNA-dependent RNA polymerase (RdRp) and, in some cases, an additional viral phosphoprotein, and they bind to, coat, and condense the viral genomic RNA. The multivalent interactions between antiterminator proteins and the viral RNA drive phase separation of the viral genome and promote replication organelle biogenesis and viral RNA replication.

Similar to replication organelle biogenesis, accumulating evidence suggests that genome packaging or nucleocapsid assembly is also mediated by phase separation, through the action of viral nucleocapsid proteins. The term ‘nucleocapsid’ refers to the nucleic acid genome surrounded by the protein coat (or capsid) of the virus. Nucleocapsid proteins, also known as coat or capsid proteins, are the viral proteins responsible for binding to, and condensing, the viral genome inside the virion.

Fascinatingly, and in further reinforcement of their parallel roles and activities, the antiterminator, phosphoprotein, and nucleocapsid functions in some viruses are performed by the same viral protein, with examples in both positive- and negative-sense RNA viruses. While these functions are often provided by the action of the nucleoprotein in negative-sense RNA viruses, these roles in positive-sense and double-stranded RNA viruses are generally separated across two (or more) viral proteins. Whether this separation of function is a result of convergent or divergent evolution remains an open question.

Importantly, viruses are simplified systems that have facilitated foundational discoveries in cell and molecular biology, including DNA replication, splicing, RNA polymerase II promoters, and oncogenes (among others). Thus, we hypothesize that studying the antiterminator, phosphoprotein, and nucleocapsid proteins of viruses may help reveal insight into the mechanistic and functional understanding of biomolecular condensates and reveal novel therapeutic strategies to tackle both viral-derived and non-communicable diseases associated with aberrant phase separation. To explore this further, we are using bioinformatic tools and in vitro liquid-liquid phase separation assays to explore the molecular grammar of these viral phase separation proteins.