The Stokes radius characteristics of reduced and carboxamidated ribonuclease A (RCAM RNase) were determined for transfer of this ''random coil'' protein from water to 1 M concentrations of the naturally occurring protecting osmolytes trimethylamine N-oxide, sarcosine, sucrose, and proline and the nonprotecting osmolyte urea. The denatured ensemble of RCAM RNase expands in urea and contracts in protecting osmolytes to extents proportional to the transfer Gibbs energy of the protein from water to osmolyte. This proportionality suggests that the sum of the transfer Gibbs energies of individual parts of the protein is responsible for the dimensional changes in the denatured ensemble. The dominant term in the transfer Gibbs energy of RCAM RNase from water to protecting osmolytes is the unfavorable interaction of the osmolyte with the peptide backbone, whereas the favorable interaction of urea with the backbone dominates in RCAM RNase transfer to urea. The side chains collectively favor transfer to the osmolytes, with some protecting osmolytes solubilizing hydrophobic side chains as well as urea does, a result suggesting there is nothing special about the ability of urea to solubilize hydrophobic groups. Protecting osmolytes stabilize proteins by raising the chemical potential of the denatured ensemble, and the uniform thermodynamic force acting on the peptide backbone causes the collateral effect of contracting the denatured ensemble. The contraction decreases the conformational entropy of the denatured state while increasing the density of hydrophobic groups, two effects that also contribute to the ability of protecting osmolytes to force proteins to fold.The adaptation of certain higher organisms to harsh environments is enabled by the intracellular presence of small organic solutes (osmolytes) that protect proteins and other cell components from the denaturing environmental stresses (1). Arakawa and Timasheff (2) have shown that the osmolytes act by raising the chemical potential of the denatured state relative to the native state, thereby increasing the (positive) Gibbs energy difference (⌬G) between the native and denatured ensembles. A pictorial description of the results of Arakawa and Timasheff is presented in the Gibbs energy diagram below, where ⌬G 1 is the unfolding Gibbs energy difference between native (N aq ) and unfolded (U aq ) protein in aqueous buffer solution, and ⌬G 3 is the Gibbs energy change for the same reaction in the presence of osmolytes. Transfer of N aq or U aq from water to osmolyte solution (N os and U os respectively) raises the chemical potential of the unfolded ensemble (⌬G 2 ) much more than it does that of the native state ensemble (⌬G 4 ), resulting in a greater stability of the protein in the osmolyte solution than in buffer, i.e., ⌬G 3 Ͼ⌬G 1 . We have determined the transfer Gibbs energy changes of amino acid side chains and peptide backbone from water to solutions of the osmolytes trimethylamine N-oxide (TMAO), sarcosine, and sucrose (3, 4). From knowledge of these tran...

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Cell fate decisions are critical for life, yet little is known about how their reliability is achieved when signals are noisy and fluctuating with time. In this study, we show that in budding yeast, the decision of cell cycle commitment (Start) is determined by the time integration of its triggering signal Cln3. We further identify the Start repressor, Whi5, as the integrator. The instantaneous kinase activity of Cln3-Cdk1 is recorded over time on the phosphorylated Whi5, and the decision is made only when phosphorylated Whi5 reaches a threshold. Cells adjust the threshold by modulating Whi5 concentration in different nutrient conditions to coordinate growth and division. Our work shows that the strategy of signal integration, which was previously found in decision-making behaviors of animals, is adopted at the cellular level to reduce noise and minimize uncertainty.DOI: http://dx.doi.org/10.7554/eLife.03977.001

Remodeling of auxin distribution during the integration of plant growth responses with the environment requires the precise control of auxin influx and efflux transporters. The plasma membrane-localized PIN-FORMED (PIN) proteins facilitate auxin efflux from cells, and their activity is regulated by reversible phosphorylation. How PIN modulates plant cellular responses to external stresses and whether its activity is coordinated by phospholipids remain unclear. Here, we reveal that, in Arabidopsis (Arabidopsis thaliana), the phosphatidic acid (PA)-regulated PINOID (PID) kinase is a crucial modulator of PIN2 activity and auxin redistribution in response to salt stress. Under salt stress, loss of phospholipase D function impaired auxin redistribution and resulted in markedly reduced primary root growth; these effects were reversed by exogenous PA. The phospholipase D-derived PA interacted with PID and increased PID-dependent phosphorylation of PIN2, which activated auxin efflux and altered auxin accumulation, promoting root growth when exposed to salt stress. Ablation of the PA binding motif not only diminished PID accumulation at the plasma membrane but also abolished PA-promoted PID phosphorylation of PIN2 and its function in coping with salt stress; however, this ablation did not affect inflorescence and cotyledon development or PIN2-dependent gravitropic and halotropic responses. Our data indicate a role for PA in coupling extracellular salt signaling to PID-directed PIN2 phosphorylation and polar auxin transport, highlighting the importance of lipid-protein interactions in the spatiotemporal regulation of auxin signaling.