Water transport in intact yeast cells as assessed by fluorescence self-quenching

Abstract

Intact yeast cells loaded with 5- and-6-carboxyfluorescein were used to assess water transport. The results were similar to those previously reported for protoplasts assessed by using either fluorescence or light scattering, and the activation energies were 8.0 and 15.1 kcal mol(-1) (33.4 and 63.2 kJ mol(-1)) for a strain overexpressing AQY1 aquaporin and a parental strain, respectively.

FIG. 1.

Equilibrium volumes of intact cells loaded with CFDA and subjected to hypo- and hyperosmotic shocks, based on a spherical cell shape. The average diameter was calculated by using the maximum and minimum dimensions of each cell (n = 90 for each data point observed by epifluorescence).

FIG. 2.

Linear relationships between Λ and ΔF and between Λ and the inverse of the relative cell volume (V₀/V). These relationships were used to calibrate the stopped-flow signals.

FIG. 3.

Fluorescence signals and the corresponding single exponential fits obtained for the overexpressing AQY1 strain loaded with CFDA subjected to hypo-, iso-, and hyperosmotic shocks at 23°C (296 K).

FIG. 4.

Arrhenius plots for the parental strain and the overexpressing AQY1 strain loaded with CFDA and subjected to a hypoosmotic shock (Λ = 0.75; temperature range, 7°C [280 K] to 38°C [311 K]). The E_a values (15.1 and 8.0 kcal mol⁻¹ [63.2 and 33.4 kJ mol⁻¹]) were calculated for the parental strain and the strain overexpressing AQY1.