Competitive intra- and extracellular nutrient sensing by the transporter homologue Ssy1p - PubMed (original) (raw)

Competitive intra- and extracellular nutrient sensing by the transporter homologue Ssy1p

Boqian Wu et al. J Cell Biol. 2006.

Abstract

Recent studies of Saccharomyces cerevisiae revealed sensors that detect extracellular amino acids (Ssy1p) or glucose (Snf3p and Rgt2p) and are evolutionarily related to the transporters of these nutrients. An intriguing question is whether the evolutionary transformation of transporters into nontransporting sensors reflects a homeostatic capability of transporter-like sensors that could not be easily attained by other types of sensors. We previously found SSY1 mutants with an increased basal level of signaling and increased apparent affinity to sensed extracellular amino acids. On this basis, we propose and test a general model for transporter- like sensors in which occupation of a single, central ligand binding site increases the activation energy needed for the conformational shift between an outward-facing, signaling conformation and an inward-facing, nonsignaling conformation. As predicted, intracellular leucine accumulation competitively inhibits sensing of extracellular amino acids. Thus, a single sensor allows the cell to respond to changes in nutrient availability through detection of the relative concentrations of intra- and extracellular ligand.

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Figures

Figure 1.

Figure 1.

Model for transporter-like sensors. States corresponding to those of a canonical transporter are presented. The elliptic object symbolizes the ligand (i.e., amino acid in the case of Ssy1p), and the vertical shaded bars indicate the membrane lipids. The dotted arrows for reaction 4 indicate that for a sensor working as proposed for Ssy1p, states O·L and I·L cannot be directly turned into one another. For a real transporter, on the other hand, reaction 4 must be efficient. The outward-facing conformation of the sensor (states O and O·L) is hypothesized to be signaling, whereas states I·L and I are nonsignaling. O, outward facing, O·L, outward facing, ligand bound; I·L, inward facing, ligand bound; I, inward facing.

Figure 2.

Figure 2.

Cytoplasmic leucine inhibits extracellular amino acid signaling by influencing the median effective concentration of extracellular amino acid. A representative experiment is shown. (A) Western blot of protein extracts of yeast cells (strain M5447) grown in minimal medium (SD) and exposed to leucine, showing processed (P) and unprocessed (U) forms of the transcription factor Stp1p. (B) Signaling measured as Stp1p processing shown in A was fitted as described (see Material and methods). Squares indicate data from A (solid curve is best fit, giving EC 50 = 12 μM), whereas triangles and broken curve indicate data and fit obtained with cells grown in SD medium with 1 mM leucine and subsequently washed (EC 50 = 66 μM).

Figure 3.

Figure 3.

Relationship between amount of cytoplasmic leucine and _EC_50 for sensing of extracellular leucine. The 23 pairs of data include experiments 1–9 presented in Table I and were fitted to the linear relationship EC 50 = _M_[Leuc] + N. Each point represents a separate condition or strain.

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