Proton- and sodium-coupled phosphate transport systems and energy status of Yarrowia lipolytica cells grown in acidic and alkaline conditions.

Abstract

In this study we have used a newly isolated Yarrowia lipolytica yeast strain with a unique capacity to grow over a wide pH range (3.5-10.5), which makes it an excellent model system for studying H(+)- and Na(+)-coupled phosphate transport systems. Even at extreme growth conditions (low concentrations of extracellular phosphate, alkaline pH values) Y. lipolytica preserved tightly-coupled mitochondria with the fully competent respiratory chain containing three points of energy conservation. This was demonstrated for the first time for cells grown at pH 9.5-10.0. In cells grown at pH 4.5, inorganic phosphate (P(i)) was accumulated by two kinetically discrete H(+)/P(i)-cotransport systems. The low-affinity system is most likely constitutively expressed and operates at high P(i) concentrations. The high-affinity system, subjected to regulation by both extracellular P(i) availability and intracellular polyphosphate stores, is mobilized during P(i)-starvation. In cells grown at pH 9.5-10, P(i) uptake is mediated by several kinetically discrete Na(+)-dependent systems that are specifically activated by Na(+) ions and insensitive to the protonophore CCCP. One of these, a low-affinity transporter operative at high P(i) concentrations is kinetically characterized here for the first time. The other two, high-affinity, high-capacity systems, are derepressible and functional during P(i)-starvation and appear to be controlled by extracellular P(i). They represent the first examples of high-capacity, Na(+)-driven P(i) transport systems in an organism belonging to neither the animal nor bacterial kingdoms. The contribution of the H(+)- and Na(+)-coupled P(i) transport systems in Y. lipolytica cells grown at different pH values was quantified. In cells grown at pH values of 4.5 and 6.0, the H(+)-coupled P(i) transport systems are predominant. The contribution of the Na(+)/P(i) cotransport systems to the total cellular P(i) uptake activity is progressively increased with increasing pH, reaching its maximum at pH 9 and higher.