Intrinsically disordered protein biosensor tracks the physical-chemical effects of osmotic stress on cells

Abstract

Cell homeostasis is perturbed when dramatic shifts in the external environment cause the physical-chemical properties inside the cell to change. Experimental approaches for dynamically monitoring these intracellular effects are currently lacking. Here, we leverage the environmental sensitivity and structural plasticity of intrinsically disordered protein regions (IDRs) to develop a FRET biosensor capable of monitoring rapid intracellular changes caused by osmotic stress. The biosensor, named SED1, utilizes the Arabidopsis intrinsically disordered AtLEA4-5 protein expressed in plants under water deficit. Computational modeling and in vitro studies reveal that SED1 is highly sensitive to macromolecular crowding. SED1 exhibits large and near-linear osmolarity-dependent changes in FRET inside living bacteria, yeast, plant, and human cells, demonstrating the broad utility of this tool for studying water-associated stress. This study demonstrates the remarkable ability of IDRs to sense the cellular environment across the tree of life and provides a blueprint for their use as environmentally-responsive molecular tools.

Competing interactions give rise to two-state behavior and switch-like transitions in charge-rich intrinsically disordered proteins

“…Precedent for switch-like transitions and dynamics that span a spectrum of timescales has been reported for different IDRs ( 10 , 13 ). Such transitions, especially in the context of tethered systems, are likely to make important contributions to the spatial and temporal control over biochemical reactions where IDRs modulate the effective concentrations of ligands around binding sites or of substrates around active sites of enzymes ( 31 , 32 , 34 , 57 , 101 – 105 ). While our results are presented for IDRs studied as autonomous units, we expect that our findings will be transferable to tethered IDRs, albeit with context-dependent modifications ( 103 , 106 ).…”

“…Studies over the past decade have helped quantify relationships ( 6 ) that connect sequence-encoded information within IDRs to properties of conformational ensembles such as overall sizes and shapes, the amplitudes of spontaneous conformational fluctuations, and the dynamics of interconverting between distinct conformational states ( 7 – 19 ). These sequence–ensemble relationships have direct functional consequences that have been uncovered via studies based on biophysical, biochemical, and engineering approaches ( 5 , 20 – 38 ). Our work, which is focused on physical principles underlying sequence–ensemble relationships of IDRs, is of direct relevance to understanding how IDRs function.…”

Phase transitions of a SEC14-like condensate at Arabidopsis plasma membranes regulate root growth

“…We further observed that during imaging, “SFH8 IDR ” puncta lacked colocalization with vesicular markers ( Figure S4F, G ), and showed LLPS hallmarks such as dynamic morphology with frequent splitting, fusion, and interconnections ( Movie S4, Figure S4H ). FRAP of mNeon-tagged SFH8 IDR puncta showed a rapid signal recovery (t1/2∼10 s, mobile fraction 40%) and sensitivity to 1,6-hexanediol which may block LLPS ( Figure S4H, I ) (Cuevas-Velazquez et al, 2021; Kroschwald et al, 2017); 1,6-hexanediol led also to the dissolution of SFH8 clusters on the PM but not of filaments ( Figure S4I ).…”

Competing interactions give rise to two-state behavior and switch-like transitions in charge-rich intrinsically disordered proteins

“…Precedent for switch-like transitions and dynamics that span a spectrum of timescales has been reported for different IDRs (10,13). Such transitions, especially in the context of tethered systems, are likely to make important contributions to the spatial and temporal control over biochemical reactions where IDRs modulate the effective concentrations of ligands around binding sites or of substrates around active sites of enzymes (31,32,34,57,(101)(102)(103)(104)(105). While our results are presented for IDRs studied as autonomous units, we expect that our findings will be transferable to tethered IDRs, albeit with context-dependent modifications (103,106).…”

“…Studies over the past decade have helped quantify relationships (6) that connect sequence-encoded information within IDRs to properties of conformational ensembles such as overall sizes and shapes, the amplitudes of spontaneous conformational fluctuations, and the dynamics of interconverting between distinct conformational states (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). These sequence-ensemble relationships have direct functional consequences that have been uncovered via studies based on biophysical, biochemical, and engineering approaches (5,(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38). Our work, which is focused on physical principles underlying sequence-ensemble relationships of IDRs, is of direct relevance to understanding how IDRs function.…”