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Abstract hnRNPA2 is a major component of mRNA transport granules in oligodendrocytes and neurons. However, the structural details of how hnRNPA2 binds the A2 recognition element (A2RE) and if this sequence stimulates granule formation by enhancing phase separation of hnRNPA2 has not yet been studied. Using solution NMR and biophysical studies, we find that each of the two individual RRMs retain the domain structure observed in complex with RNA but are not rigidly confined (i.e. they move independently) in solution in the absence of RNA. hnRNPA2 RRMs bind the minimal rA2RE11 weakly but at least, and most likely, two hnRNPA2 molecules are able to simultaneously bind the longer 21mer myelin basic protein A2RE. Upon binding of the RNA, NMR chemical shift deviations are observed in both RRMs, suggesting both play a role in binding the A2RE11. Interestingly, addition of short A2RE RNAs or longer RNAs containing this sequence completely prevents in vitro phase separation of full-length hnRNPA2 and aggregation of the disease-associated mutants. These findings suggest that RRM interactions with specific recognition sequences alone do not account for nucleating granule formation, consistent with models where multivalent protein:RNA and protein:protein contacts form across many sites in granule proteins and long RNA transcripts. 

Membrane bending is a ubiquitous cellular process that is required for membrane traffic, cell motility, organelle biogenesis, and cell division. Proteins that bind to membranes using specific structural features, such as wedge-like amphipathic helices and crescent-shaped scaffolds, are thought to be the primary drivers of membrane bending. However, many membrane-binding proteins have substantial regions of intrinsic disorder which lack a stable three-dimensional structure. Interestingly, many of these disordered domains have recently been found to form networks stabilized by weak, multivalent contacts, leading to assembly of protein liquid phases on membrane surfaces. Here we ask how membrane-associated protein liquids impact membrane curvature. We find that protein phase separation on the surfaces of synthetic and cell-derived membrane vesicles creates a substantial compressive stress in the plane of the membrane. This stress drives the membrane to bend inward, creating protein-lined membrane tubules. A simple mechanical model of this process accurately predicts the experimentally measured relationship between the rigidity of the membrane and the diameter of the membrane tubules. Discovery of this mechanism, which may be relevant to a broad range of cellular protrusions, illustrates that membrane remodeling is not exclusive to structured scaffolds but can also be driven by the rapidly emerging class of liquid-like protein networks that assemble at membranes.