Pressure Sensitivity of SynGAP/PSD‐95 Condensates as a Model for Postsynaptic Densities and Its Biophysical and Neurological Ramifications

Abstract

Biomolecular condensates consisting of proteins and nucleic acids can serve critical biological functions, so that some condensates are referred as membraneless organelles. They can also be disease‐causing, if their assembly is misregulated. A major physicochemical basis of the formation of biomolecular condensates is liquid–liquid phase separation (LLPS). In general, LLPS depends on environmental variables, such as temperature and hydrostatic pressure. The effects of pressure on the LLPS of a binary SynGAP/PSD‐95 protein system mimicking postsynaptic densities, which are protein assemblies underneath the plasma membrane of excitatory synapses, were investigated. Quite unexpectedly, the model system LLPS is much more sensitive to pressure than the folded states of typical globular proteins. Phase‐separated droplets of SynGAP/PSD‐95 were found to dissolve into a homogeneous solution already at ten‐to‐hundred bar levels. The pressure sensitivity of SynGAP/PSD‐95 is seen here as a consequence of both pressure‐dependent multivalent interaction strength and void volume effects. Considering that organisms in the deep sea are under pressures up to about 1 kbar, this implies that deep‐sea organisms have to devise means to counteract this high pressure sensitivity of biomolecular condensates to avoid harm. Intriguingly, these findings may shed light on the biophysical underpinning of pressure‐related neurological disorders in terrestrial vertebrates.

Comparative roles of charge, π , and hydrophobic interactions in sequence-dependent phase separation of intrinsically disordered proteins

“…By elucidating physical aspects of the residueresidue contacts in condensates, highlighting these contacts' largely solvent-exposed character with ramifications such as reduced hydrophobic strengths relative to buried nonpolar contacts, fundamental conceptual and quantitative progress is made toward predictive models for biomolecular condensates. (20,43,51,54), hydrostatic pressure (62)(63)(64), salt (41,46), and osmolyte (27,63), offering physical insights into the LLPS behaviors of, for example, the DEAD-box RNA helicase Ddx4 (34,65), RNA-binding protein fused in sarcoma (FUS) (52), prion-like domains (60), and postsynaptic densities (64).…”

Protein folding and surface interaction phase diagrams in vitro and in cells

“…Thus, LLPSRs can regulate binding and signaling by affecting accessibility of protein surfaces. Phase-separated condensates of SynGAP and PSD-95, important for synaptic communication, are also very sensitive to pressure and could play a role in decompression sickness [63], when divers return from a high-pressure environment.…”

Assembly of Model Postsynaptic Densities Involves Interactions Auxiliary to Stoichiometric Binding

“…These include RNA binding proteins [52], transmembrane layer proteins such as N-methyl D-aspartate receptor (NMDAR) [53], actins in the cytoskeleton layer, scaffold proteins such as PDZ-domaincontaining PSD-95s, which are membrane-associated guanylate kinases (MAGUKs) [54], guanylate-kinase (GK) associated protein (GKAP) [55], SH3 and multiple ankyrin repeat domain proteins (Shanks) [56], and Homer family of adaptor proteins [57], as well as the naptic membranes, PSDs can exchange materials with the cytoplasm in synaptic spines and, accordingly, serves to provide spatial and temporal organization of the neurotransmitter receptors at the synapse [43]. The functional importance of PSDs is highlighted by intriguing recent observations that PSDs are downsized during sleep [59,60], which suggest that synaptic strength is renormalized during sleep; and that a SynGAP-PSD-95 condensate model of PSD [42] can be disassembled by moderate hydrostatic pressure [16,61], which points to a possible biophysical origin of the high pressure neurological syndrome experienced by divers [62].…”

“…Subsequently, more realistic in vitro models of PSD, encompassing SynGAP and three scaffold proteins GKAP, Shank3, and Homer3 (in addition to PSD-95) as well as NR2B as another glutamate receptor, have also been constructed as a versatile research platform [8]. Here, as a first step toward elucidating the statistical mechanics of PSD assembly, we focus on the simpler two-component SynGAP-PSD-95 construct, the same system we have utilized recently for studying the effect of hydrostatic pressure on PSD stability [61]. SynGAP and PSD-95 undergo LLPS together when mixed but each of the individual components, SynGAP or PSD-95 (up to 100 µM concentration each), does not phase separate by itself [42].…”