Chloroplast biogenesis. Cell-free transfer of envelope monogalactosylglycerides to thylakoids

Abstract

An ATP- and temperature-dependent transfer of monogalactosylglycerides from the chloroplast envelope to the chloroplast thylakoids was reconstituted in a cell-free system prepared from isolated chloroplasts of garden pea ( Pisum sativum) or spinach ( Spinacia oleracea). Isolated envelope membranes, in which the label was present exclusively in monogalactosylglycerides, were prepared radiolabeled in vitro with [ 14C]galactose from UDP-[ 14C]galactose to label galactolipids as the donor. ATP-dependent transfer of radioactivity from donor to unlabeled acceptor thylakoids, immobilized on nitrocellulose strips, was observed. In some experiments linear transfer for longer than 30 min of incubation was facilitated by the addition of stroma proteins but in other experiments stroma was without effect or inhibitory suggesting no absolute requirements for a soluble protein carrier. Transfer was donor specific. No membrane fraction tested (plasma membrane, tonoplast, endoplasmic reticulum, nuclei, Golgi apparatus, mitochondria or thylakoids) (isolated from tissue radiolabeled in vivo with [ 14C]acetate) other than chloroplast envelopes demonstrated any significant ability to transfer labeled membrane lipids to immobilized thylakoids. Acceptor specificity, while not absolute, showed a 3–10-fold greater ATP-dependent transfer of labeled galactolipids from chloroplast envelopes to immobilized thylakoids than to other leaf membranes. The results provide independent confirmation of the potential for transfer of galactolipids between chloroplast envelopes and thylakoids suggested previously from ultrastructural studies and of the known location of thylakoid galactolipid biosynthetic activities in the chloroplast envelope.