Molecular interactions contributing to FUS SYGQ LC-RGG phase separation and co-partitioning with RNA polymerase II heptads

The RNA-binding protein FUS (Fused in Sarcoma) mediates phase separation in biomolecular condensates and functions in transcription by clustering with RNA polymerase II. Specific contact residues and interaction modes formed by FUS and the C-terminal heptad repeats of RNA polymerase II (CTD) have been suggested but not probed directly. Here we show how RGG domains contribute to phase separation with the FUS N-terminal low-complexity domain (SYGQ LC) and RNA polymerase II CTD. Using NMR spectroscopy and molecular simulations, we demonstrate that many residue types, not solely arginine-tyrosine pairs, form condensed-phase contacts via several interaction modes including, but not only sp 2 - π and cation- π interactions. In phases also containing RNA polymerase II CTD, many residue types form contacts, including both cation- π and hydrogen-bonding interactions formed by the conserved human CTD lysines. Hence, our data suggest a surprisingly broad array of residue types and modes explain co-phase separation of FUS and RNA polymerase II. NMR visualization of phase-separated FUS and RNA polymerase II domains in models of transcriptional condensates show that a much wider array of residue types and interaction modes stabilize phases than previously proposed.