ABSTRACT
Chloroplasts and mitochondria are unique endosymbiotic cellular organelles surrounded by two membranes. Essential metabolic networking between these compartments and their hosting cells requires the exchange of a large number of biochemical pathway intermediates in a directed and coordinated fashion across their inner and outer envelope membranes. Here, we describe the identification and functional characterization of a highly specific, regulated solute channel in the outer envelope of chloroplasts, named OEP40. Loss of OEP40 function in Arabidopsis thaliana results in early flowering under cold temperature. The reconstituted recombinant OEP40 protein forms a high conductance ß-barrel ion channel with subconductant states in planar lipid bilayers. The OEP40 channel is slightly cation-selective PK+/PCl- ≈ 4:1 and rectifying (iâ/iâ â 2) with a slope conductance of Gmax â 690 picosiemens. The OEP40 channel has a restriction zone diameter of â 1.4 nm and is permeable for glucose, glucose 1-phosphate and glucose 6-phosphate, but not for maltose. Moreover, channel properties are regulated by trehalose 6-phosphate, which cannot permeate. Altogether, our results indicate that OEP40 is a "glucose-gate" in the outer envelope membrane of chloroplasts, facilitating selective metabolite exchange between chloroplasts and the surrounding cell.
Subject(s)
Arabidopsis Proteins/chemistry , Arabidopsis/chemistry , Chloroplast Proteins/chemistry , Chloroplasts/chemistry , Intracellular Membranes/chemistry , Membrane Proteins/chemistry , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Chloroplast Proteins/genetics , Chloroplast Proteins/metabolism , Chloroplasts/metabolism , Glucose/chemistry , Glucose/genetics , Glucose/metabolism , Intracellular Membranes/metabolism , Lipid Bilayers/chemistry , Lipid Bilayers/metabolism , Membrane Proteins/genetics , Membrane Proteins/metabolismABSTRACT
BACKGROUND: Metabolite, ion and protein translocation into chloroplasts occurs across two membranes, the inner and the outer envelope. Solute and metabolite channels fulfill very important functions in integrating the organelles into the metabolic network of the cell. However so far only a few have been identified. Here we describe the identification and the characterization of the outer envelope protein of 23 kDa, Oep23 from garden pea. RESULTS: Oep23 is found in the entire plant lineage from green algae to flowering plants. It is expressed in all organs and developmental states tested so far. The reconstituted recombinant protein Oep23 from pea forms a high conductance ion channel with a maximal conductance in the fully open state of 466 ± 14pS at a holding potential of +100 mV (in 250 mM KCl). The Oep23 channel is cation selective (PK+ : PCl- = 15 : 1) with a voltage dependent open probability of maximal Vmem = 0 mV. CONCLUSION: The data indicate that the Oep23 activity represents a single channel unit and does not assemble into a multiple pore complex like bacterial type porins or mitochondrial voltage dependent anion channel. Thus, Oep23 represents a new member of ion channels in the outer envelope of chloroplasts involved in solute exchange.
