A rectifying ATP-regulated solute channel in the chloroplastic outer envelope from pea

Abstract

B.Bölter and R.Hemmler contributed equally to this workPhosphorylated carbohydrates are the main photoassimilated export products from chloroplasts that support the energy household and metabolism of the plant cell. Channels formed by the chloroplastic outer envelope protein OEP21 selectively facilitate the translocation of triosephosphate, 3-phosphoglycerate and phosphate, central intermediates in the source-sink relationship between the chloroplast and the cytosol. The anion selectivity and asymmetric transport properties of OEP21 are modulated by the ratio between ATP and triosephosphates, 3-phosphoglycerate and phosphate in the intermembrane space. Conditions that lead to export of triosephosphate from chloroplasts, i.e. photosynthesis, result in outward-rectifying OEP21 channels, while a high ATP to triosephosphate ratio, e.g. dark metabolism, leads to inward-rectifying OEP21 channels with a less pronounced anion selectivity. We conclude that solute exchange between plastids and cytosol can already be regulated at the level of the organellar outer membrane.

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“…The painting technique was used to form planar lipid bilayers 5,16 . The cis chamber solution was changed to asymmetrical concentrations (cis chamber: 250 mM KCl, 1 mM CaCl 2 and 10 mM MOPS-Tris, pH 7.0) once a stable bilayer was formed in symmetrical solutions of 20 mM KCl and 10 mM MOPS-Tris, pH 7.0.…”

“…The tim23-2 mitochondria specifically processed and imported the protein in a membrane potentialdependent manner (Fig. 5a, lanes [16][17][18][19][20] with an efficiency of 50-70% relative to wild type mitochondria. Similar import efficiencies were seen with purified inner membrane vesicles (not shown).…”

Insertion of OEP14 into the Outer Envelope Membrane Is Mediated by Proteinaceous Components of Chloroplasts

“…The second group of proteins consists of most of the outer membrane proteins identified so far. These outer membrane proteins are synthesized at their mature size in the cytosol without a cleavable transit peptide (Salomon et al, 1990;Li et al, 1991;Ko et al, 1992;Fischer et al, 1994;Kessler et al, 1994;Seedorf et al, 1995;Bolter et al, 1999). The import mechanism for the first group of proteins has been studied extensively, and many protein components of the import machinery have been identified (Chen and Schnell, 1999;Keegstra and Cline, 1999;May and Soll, 1999).…”

A Proteomics Dissection of Arabidopsis thaliana Vacuoles Isolated from Cell Culture

“…Fifteen micrograms of proteins from protoplast or vacuole extracts were loaded on a 12% SDS-polyacrylamide gel and, after migration at 200 V, transferred to a nitrocellulose membrane. The presence of different characteristic proteins was assessed using Western blot analyses and primary polyclonal antibodies raised against the outer envelop protein 21 (45) and the light-harvesting complex b (O. Vallon, Institut de Biologie Physico-Chimique, Paris, France) (plastid markers), the preprotein translocase of the mitochondrial outer membrane (TOM40, channel-forming subunit (46)), the plasma membrane P-type H ϩ -ATPase (PMA2 (47)), and the HDEL domain of the endoplasmic reticulum (ER) proteins (48,49). Antibodies raised against tonoplastic markers such as the tobacco ␣-TIP (50) and the cauliflower ␥-TIP (51) were used to control for vacuole enrichment.…”