Albert-Jean Dorne

Preparation and Characterization of Membrane Fractions Enriched in Outer and Inner Envelope Membranes from Spinach Chloroplasts

A gentle osmotic shock of intact and purified chloroplasts allows the preparation of envelope membranes in a reasonably pure state. Unfortunately, irreversible changes take place in the membranes (fusions?) during the osmotic shock thus making the separation of the outer membrane from the inner impossible (Douce, Joyard, 1979). Such a separation can only be achieved by strikingly different procedures. Cline et al. (1981) have described a procedure which includes freeze-thaw lysis of intact pea chloroplasts. We have developed a different method to provide the separation of two membrane fractions deriving from the chloroplast envelope membranes. Since we have characterized two polypeptides as outer envelope polypeptides (Joyard et al., 1983), we were able to characterize the membrane fractions obtained (Block et al., in press).

The galactolipid:galactolipid galactosyltransferase is located on the outer surface of the outer membrane of the chloroplast envelope

1. INTRODUCTION It is well known that the plastid envelope is in- volved in the synthesis of galactolipids (for a re- view see [l]). For instance it has been demon- strated that at least two distinct enzymes res- ponsible for the synthesis of monogalactosyldiacyl- glycerol (MGDG) and digalactosyldiacylglycerol (DGDG) are associated with chloroplast envelope membranes. The first enzyme, UDP-galactose:di- acylglycerol galactosyltransferase, catalyses the in- corporation of galactose from UDP-galactose into MGDG [3-51:

The Plastid Envelope Membranes: Their Structure, Composition, and Role in Chloroplast Biogenesis

The feature shared by all members of the plastid family (proplastids, elaioplasts, leukoplasts, amyloplasts, chromoplasts, etioplasts, and chloroplasts) is a pair of outer membranes, known as the envelope. These membranes together provide a flexible boundary between the plastid and the surrounding cytosol (Thomson, 1974). This applies even to highly senescent plastids (Priestley, 1977), and to plastids devoid of ribosomes (Borner et al., 1976; Feierabend and Schrader-Reichhardt, 1976)—from which all other membraneous structures have disappeared—and to dividing plastids (Chaly et al.,1980). Consequently, the plastid envelope is a permanent structure in the sense that every existing envelope membrane could theoretically be traced back through generations of cells and organisms maintaining uninterruptedly for millions of years some specific structural organization.

The acyl-CoA synthetase and acyl-CoA thioesterase are located on the outer and inner membrane of the chloroplast envelope, respectively

The chloroplast envelope contains an acyl‐CoA synthetase and an acyl‐CoA thioesterase which are associated with the outer and inner membrane, respectively.

Pea Seed Mitochondria Are Endowed with a Remarkable Tolerance to Extreme Physiological Temperatures

Most seeds are anhydrobiotes, relying on an array of protective and repair mechanisms, and seed mitochondria have previously been shown to harbor stress proteins probably involved in desiccation tolerance. Since temperature stress is a major issue for germinating seeds, the temperature response of pea (Pisum sativum) seed mitochondria was examined in comparison with that of mitochondria from etiolated epicotyl, a desiccation-sensitive tissue. The functional analysis illustrated the remarkable temperature tolerance of seed mitochondria in response to both cold and heat stress. The mitochondria maintained a well-coupled respiration between −3.5°C and 40°C, while epicotyl mitochondria were not efficient below 0°C and collapsed above 30°C. Both mitochondria exhibited a similar Arrhenius break temperature at 7°C, although they differed in phospholipid composition. Seed mitochondria had a lower phosphatidylethanolamine-to-phosphatidylcholine ratio, fewer unsaturated fatty acids, and appeared less susceptible to lipid peroxidation. They also accumulated large amounts of heat shock protein HSP22 and late-embryogenesis abundant protein PsLEAm. The combination of membrane composition and stress protein accumulation required for desiccation tolerance is expected to lead to an unusually wide temperature tolerance, contributing to the fitness of germinating seeds in adverse conditions. The unique oxidation of external NADH at low temperatures found with several types of mitochondria may play a central role in maintaining energy homeostasis during cold shock, a situation often encountered by sessile and ectothermic higher plants.

The phosphatidic acid phosphatase of the chloroplast envelope is located on the inner envelope membrane

The envelope from spinach chloroplasts contains an alkaline phosphatidic acid phosphatase which was found to be located on the inner envelope membrane. The diacylglycerol formed by this enzyme from endogenous phosphatidic acid is then used as a substrate for galactolipid synthesis on the inner envelope membrane.

Protein-mediated transfer of phosphatidylcholine from liposomes to spinach chloroplast envelope membranes

Abstract We have demonstrated that an active transfer of phosphatidylcholine from liposomes towards spinach chloroplast envelope was catalyzed by a phospholipid-transfer protein purified from spinach leaves. The transfer is actually a complex process. During the first 10 min of the incubation, the exchange of phosphatidylcholine between liposomes and isolated envelope vesicles was predominant, as shown by the equilibration of phosphatidylcholine specific activity to the same level in both the liposomes and the envelope vesicles. Further incubation led to a 35% increase of the phosphatidylcholine content of envelope membranes, thus corresponding to a net transfer of phosphatidylcholine from liposomes towards envelope vesicles. After incubation of intact chloroplasts and liposomes in the presence of purified phospholipid-transfer protein, most of the radioactive phosphatidylcholine transferred to intact chloroplasts was recovered with the envelope membrane fraction. In addition, a mild phospholipase C treatment of intact chloroplasts after phosphatidylcholine transfer has demonstrated that all the radioactive phosphatidylcholine remained in the cytosolic leaflet of the outer envelope membrane and was not redistributed towards internal chloroplast membranes. Such a result, which mimics the in vivo situation, suggests that the phospholipid-transfer protein might be partly responsible (together with the apparent lack of transmembrane lipid diffusion) for the different lipid composition of the outer envelope membrane (when compared with the other plastid membranes) and for the asymmetrical distribution of phosphatidylcholine within this membrane.

Localization of galactolipid: galactolipid galactosyltransferase and acyltransferase in outer envelope membrane of spinach chloroplasts

Abstract We have measured the localization within spinach chloroplast envelope membranes of two galactolipidmanipulating enzymes, the galactolipid:galactolipid galactosyltransferase and the galactolipid:galactolipid acyltransferase. Both are localized on the outer envelope membrane. This situation differs strikingly from the localization of the enzymes involved in monogalactosyldiacylglycerol synthesis, on the inner envelope membrane.

Polar lipid composition of leaves from nine typical alpine species

Abstract The fatty acids and polar lipid compositions of leaves from nine alpine species were almost identical to that of plants growing in habitats with little seasonal variation in temperature. Furthermore each polar lipid had about the same fatty acid composition in all plant species studied. It is suggested that neither the relative proportions of different lipid classes nor the degree of saturation of individual classes are directly implicated in the adaptation of plant tissues to different climates.

Lipid Distribution and Synthesis Within the Plant Cell

The higher plant cell contains numerous distinct organelles or membranes1 but only some of these have been properly purified and characterized. Determination of the in vivo glycerolipid composition of these plant cell organelles or membranes, and their role in lipid metabolism is not simple, in contrast to what is often believed.