Title

Quantitative proteomics of the tonoplast reveals a role for glycolytic enzymes in salt tolerance

Document Type

Article

Publication details

Barkla, BJ, Vera-Estrella, R, Hernandez-Coronado, M & Pantoja, O 2009, 'Quantitative proteomics of the tonoplast reveals a role for glycolytic enzymes in salt tolerance', American Society of Plant Biologists, vol. 21, no. 12, pp. 4044-4058.

Published version available from:

http://dx.doi.org/10.1105/tpc.109.069211

Abstract

To examine the role of the tonoplast in plant salt tolerance and identify proteins involved in the regulation of transporters for vacuolar Na+ sequestration, we exploited a targeted quantitative proteomics approach. Two-dimensional differential in-gel electrophoresis analysis of free flow zonal electrophoresis separated tonoplast fractions from control, and salt-treated Mesembryanthemum crystallinum plants revealed the membrane association of glycolytic enzymes aldolase and enolase, along with subunits of the vacuolar H+-ATPase V-ATPase. Protein blot analysis confirmed coordinated salt regulation of these proteins, and chaotrope treatment indicated a strong tonoplast association. Reciprocal coimmunoprecipitation studies revealed that the glycolytic enzymes interacted with the V-ATPase subunit B VHA-B, and aldolase was shown to stimulate V-ATPase activity in vitro by increasing the affinity for ATP. To investigate a physiological role for this association, the Arabidopsis thaliana cytoplasmic enolase mutant, los2, was characterized. These plants were salt sensitive, and there was a specific reduction in enolase abundance in the tonoplast from salt-treated plants. Moreover, tonoplast isolated from mutant plants showed an impaired ability for aldolase stimulation of V-ATPase hydrolytic activity. The association of glycolytic proteins with the tonoplast may not only channel ATP to the V-ATPase, but also directly upregulate H+-pump activity.