Effects of decreased cytosolic pH on water permeability of intact yeast
protoplasts expressing native or mutant NtPIP2;1. Kinetics of protoplast
swelling was recorded as time courses of decreased scattered light intensity
(arbitrary units) in a stopped flow spectrophotometer. Yeast protoplasts
expressing no extra aquaporin exhibit very low water permeability rates
(upper three curves in a chart), whereas those expressing NtPIP2;1
completed swelling in 0.2 s following osmotic shift. A change of cytosolic pH
was achieved by incubation of yeast spheroblasts expressing unaltered NtPIP2;1
(A) or mutant NtPIP2;1-S118A (B), NtPIP2;1-S277A
(C), NtPIP2;1-H196A (D), or NtPIP2;1-H196A/S277A NtPIP2;1
(E) in buffers at pH 7.5 and pH 5, respectively, and an additional
acid load treatment with 50 mm NaAc at pH 5. (n = 30
± S.E.).