Phase transitions in the assembly of multivalent signalling proteins

Abstract

Cells are organized on length scales ranging from Angstroms to microns. However, the mechanisms by which Angstrom-scale molecular properties are translated to micron-scale macroscopic properties are not well understood. Here we show that interactions between diverse, synthetic multivalent macromolecules (including multi-domain proteins and RNA) produce sharp, liquid-liquid demixing phase separations, generating micron-sized liquid droplets in aqueous solution. This macroscopic transition corresponds to a molecular transition between small complexes and large, dynamic supramolecular polymers. The concentrations needed for phase transition are directly related to valency of the interacting species. In the case of the actin regulatory protein, neuronal Wiskott-Aldrich Syndrome Protein (N-WASP) interacting with its established biological partners Nck and phosphorylated nephrin1, the phase transition corresponds to a sharp increase in activity toward the actin nucleation factor, Arp2/3 complex. The transition is governed by the degree of phosphorylation of nephrin, explaining how this property of the system can be controlled to regulatory effect by kinases. The widespread occurrence of multivalent systems suggests that phase transitions are likely used to spatially organize and biochemically regulate information throughout biology.

Tau protein liquid–liquid phase separation can initiate tau aggregation

“…Phosphorylation introduces negative charges into proteins and changes the charge distribution and the electrostatic interactions along the polypeptide backbone, and hence also the LLPS characteristics of proteins. For example, phosphorylation regulates LLPS of nephrine (Li et al , 2012), FUS (Monahan et al , 2017), and RNA binding protein CPEB4 (Guillén‐Boixet et al , 2016). Phosphorylation is also the major type of PTMs in tau (Morris et al , 2015), and more than 80 potential phosphorylation sites can be found in the amino acid sequence of tau (Reynolds et al , 2008).…”

“…Intracellular LLPS and the resulting functional membrane‐less organelles, which biophysically can be described as gels or polyelectrolyte brushes, are emerging concepts in cell biology, which are now implicated in manifold cellular functions and applications (Li et al , 2012; Aumiller et al , 2014; Elbaum‐Garfinkle et al , 2015; Jiang et al , 2015; Aguzzi & Altmeyer, 2016; Mitrea et al , 2016; Schmidt & Görlich, 2016; Schmidt & Rohatgi, 2016). For research in neurodegenerative diseases, in particular, the function and misfunction of LLPS by involved proteins—such as tau, FUS, TDP43, hnRNPA1, C9orf72 dipeptide repeats—introduces a novel exciting concept that may provide a common underlying mechanism in these diseases.…”

Higher‐order oligomerization promotes localization of SPOP to liquid nuclear speckles

“…Membrane‐less bodies with liquid‐like properties are characterized by dynamic exchange of components with their surroundings (Li et al , 2012b; Molliex et al , 2015; Nott et al , 2015) and adopt spherical shapes (Brangwynne et al , 2011; Patel et al , 2015). When we photobleached individual nuclear speckles, their fluorescence intensity recovered with a characteristic time of 15 ± 1 s, demonstrating that protein molecules enter and leave the speckles quickly (Fig 3C).…”

“…Protein/protein and protein/RNA interactions promote assembly of and recruitment to these cellular bodies (Tourrière et al , 2003). Their liquid nature is thought to be generated via the process of liquid demixing phase separation (Brangwynne et al , 2009; Li et al , 2012b; Molliex et al , 2015; Nott et al , 2015; Patel et al , 2015), which leads to large, micrometer‐sized assemblies above threshold macromolecular concentrations. We show that nuclear speckles have liquid droplet properties and that they, together with nucleoli (Brangwynne et al , 2011), P granules (Brangwynne et al , 2009), and stress granules (Molliex et al , 2015), belong to the category of liquid membrane‐less organelles.…”

“…The inherent size heterogeneity of such higher‐order assemblies generates challenges for their structural and mechanistic characterization, but recent progress has provided insight into their ability to mediate signal amplification, filter noise, and regulate signaling in time and space (Li et al , 2012b; Wu, 2013). Some of these assemblies form via nucleation‐driven polymerization and result in stable complexes, and we are starting to understand the structural basis for their proximity‐enhanced activation (Korennykh et al , 2009; Yin et al , 2009; Li et al , 2012a; Lu et al , 2014).…”