Biomedical subjects

Christoph Benning

Publications and source records attributed to Christoph Benning.

26 records · Page 2Linked to original sources

PubMed2003

Native uridine 5'-diphosphate-sulfoquinovose synthase, SQD1, from spinach purifies as a 250-kDa complex.

Sulfoquinovosyldiacylglycerol is a polar lipid present in photosynthetic membranes. It contributes to the negative surface charge of the membrane and plays a pivotal role under phosphate stress. The SQD1 protein is the key enzyme involved in the formation of the sulfolipid head group precursor, uridine 5(')-diphosphate (UDP)-sulfoquinovose, from UDP-glucose and sulfite. A cDNA encoding the spinach SQD1 protein was isolated and functionally expressed in Escherichia coli. The recombinant enzyme was compared to the native enzyme purified from isolated spinach chloroplasts. While the K(m) for UDP-glucose was indistinguishable for the two forms, the K(m) for sulfite was more than fourfold lower (< microM) for the native enzyme. Sizing by gel filtration indicated that the native form purified as a large complex of approximately 250 kDa, which is more than twice as large as the calculated size for the homodimer. It is proposed that in vivo SQD1 forms a complex with accessory proteins.

Amino Acid Sequence↗

PubMed2003

Anionic lipids are required for chloroplast structure and function in Arabidopsis.

Photosynthetic membranes of plants primarily contain non-phosphorous glycolipids. The exception is phosphatidylglycerol (PG), which is an acidic/anionic phospholipid. A second major anionic lipid in chloroplasts is the sulfolipid sulfoquinovosyldiacylglycerol (SQDG). It is hypothesized that under severe phosphate limitation, SQDG substitutes for PG, ensuring a constant proportion of anionic lipids even under adverse conditions. A newly constructed SQDG and PG-deficient double mutant supports this hypothesis. This mutant, sqd2 pgp1-1, carries a T-DNA insertion in the structural gene for SQDG synthase (SQD2) and a point mutation in the structural gene for phosphatidylglycerolphosphate synthase (PGP1). In the sqd2 pgp1-1 double mutant, the fraction of total anionic lipids is reduced by approximately one-third, resulting in pale yellow cotyledons and leaves with reduced chlorophyll content. Photoautotrophic growth of the double mutant is severely compromised, and its photosynthetic capacity is impaired. In particular, photosynthetic electron transfer at the level of photosystem II (PSII) is affected. Besides these physiological changes, the mutant shows altered leaf structure, a reduced number of mesophyll cells, and ultrastructural changes of the chloroplasts. All observations on the sqd2 pgp1-1 mutant lead to the conclusion that the total content of anionic thylakoid lipids is limiting for chloroplast structure and function, and is critical for overall photoautotrophic growth and plant development.

Anions↗

PubMed2002

Arabidopsis disrupted in SQD2 encoding sulfolipid synthase is impaired in phosphate-limited growth.

The sulfolipid sulfoquinovosyldiacylglycerol is one of the three nonphosphorous glycolipids that provide the bulk of the structural lipids in photosynthetic membranes of seed plants. Unlike the galactolipids, sulfolipid is anionic at physiological pH because of its 6-deoxy-6-sulfonate-glucose (sulfoquinovose) head group. The biosynthesis of this lipid proceeds in two steps: first, the assembly of UDP-sulfoquinovose from UDP-glucose and sulfite, and second, the transfer of the sulfoquinovose moiety from UDP-sulfoquinovose to diacylglycerol. The first reaction is catalyzed by the SQD1 protein in Arabidopsis. Here we describe the identification of the SQD2 gene of Arabidopsis. We propose that this gene encodes the sulfoquinovosyltransferase catalyzing the second step of sulfolipid biosynthesis. Expression of SQD1 and SQD2 in Escherichia coli reconstituted plant sulfolipid biosynthesis in this bacterium. Insertion of a transfer DNA into this gene in Arabidopsis led to complete lack of sulfolipid in the respective sqd2 mutant. This mutant showed reduced growth under phosphate-limited growth conditions. The results support the hypothesis that sulfolipid can function as a substitute of anionic phospholipids under phosphate-limited growth conditions. Along with phosphatidylglycerol, sulfolipid contributes to maintaining a negatively charged lipid-water interface, which presumably is required for proper function of photosynthetic membranes.

Arabidopsis↗

PubMed2002

Galactolipids rule in seed plants.

Chloroplast membranes contain high levels of the galactolipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG). The isolation of the genes involved in the biosynthesis of MGDG and DGDG, and the identification of galactolipid-deficient Arabidopsis mutants has greatly facilitated the analysis of galactolipid biosynthesis and function. Galactolipids are found in X-ray structures of photosynthetic complexes, suggesting a direct role in photosynthesis. Furthermore, galactolipids can substitute for phospholipids, as suggested by increases in the galactolipid:phospholipid ratio after phosphate deprivation. The ratio of MGDG to DGDG is also crucial for the physical phase of thylakoid membranes and might be regulated.

Arabidopsis↗

PubMed2002

The pgp1 mutant locus of Arabidopsis encodes a phosphatidylglycerolphosphate synthase with impaired activity.

Phosphatidylglycerol is a ubiquitous phospholipid that is also present in the photosynthetic membranes of plants. Multiple independent lines of evidence suggest that this lipid plays a critical role for the proper function of photosynthetic membranes and cold acclimation. In eukaryotes, different subcellular compartments are competent for the biosynthesis of phosphatidylglycerol. Details on the plant-specific pathways in different organelles are scarce. Here, we describe a phosphatidylglycerol biosynthesis-deficient mutant of Arabidopsis, pgp1. The overall content of phosphatidylglycerol is reduced by 30%. This mutant carries a point mutation in the CDP-alcohol phosphotransferase motif of the phosphatidylglycerolphosphate synthase (EC 2.7.8.5) isoform encoded by a gene on chromosome 2. The mutant shows an 80% reduction in plastidic phosphatidylglycerolphosphate synthase activity consistent with the plastidic location of this particular isoform. Mutant plants are pale green, and their photosynthesis is impaired. This mutant provides a promising new tool to elucidate the biosynthesis and function of plastidic phosphatidylglycerol in seed plants.

Amino Acid Sequence↗

PubMed2002

Digalactosyldiacylglycerol synthesis in chloroplasts of the Arabidopsis dgd1 mutant.

Galactolipid biosynthesis in plants is highly complex. It involves multiple pathways giving rise to different molecular species. To assess the contribution of different routes of galactolipid synthesis and the role of molecular species for growth and photosynthesis, we initiated a genetic approach of analyzing double mutants of the digalactosyldiacylglycerol (DGDG) synthase mutant dgd1 with the acyltransferase mutant, act1, and the two desaturase mutants, fad2 and fad3. The double mutants showed different degrees of growth retardation: act1,dgd1 was most severely affected and growth of fad2,dgd1 was slightly reduced, whereas fad3,dgd1 plants were very similar to dgd1. In act1,dgd1, lipid and chlorophyll content were reduced and photosynthetic capacity was affected. Molecular analysis of galactolipid content, fatty acid composition, and positional distribution suggested that the growth deficiency is not caused by changes in galactolipid composition per se. Chloroplasts of dgd1 were capable of synthesizing monogalactosyldiacylglycerol, DGDG, and tri- and tetragalactosyldiacylglycerol. Therefore, the reduced growth of act1,dgd1 and fad2,dgd1 cannot be explained by the absence of DGDG synthase activity from chloroplasts. Molecular analysis of DGDG accumulating in the mutants during phosphate deprivation suggested that similarly to the residual DGDG of dgd1, this additional lipid is synthesized in association with chloroplast membranes through a pathway independent of the mutations, act1, dgd1, fad2, and fad3. Our data imply that the severe growth defect of act1,dgd1 is caused by a reduced metabolic flux of chloroplast lipid synthesis through the eukaryotic and prokaryotic pathway as well as by the reduction of photosynthetic capacity caused by the destabilization of photosynthetic complexes.

Actins↗

PubMed2002

Contrapuntal networks of gene expression during Arabidopsis seed filling.

We have used cDNA microarrays to examine changes in gene expression during Arabidopsis seed development and to compare wild-type and mutant wrinkled1 (wri1) seeds that have an 80% reduction in oil. Between 5 and 13 days after flowering, a period preceding and including the major accumulation of storage oils and proteins, approximately 35% of the genes represented on the array changed at least twofold, but a larger fraction (65%) showed little or no change in expression. Genes whose expression changed most tended to be expressed more in seeds than in other tissues. Genes related to the biosynthesis of storage components showed several distinct temporal expression patterns. For example, a number of genes encoding core fatty acid synthesis enzymes displayed a bell-shaped pattern of expression between 5 and 13 days after flowering. By contrast, the expression of storage proteins, oleosins, and other known abscisic acid-regulated genes increased later and remained high. Genes for photosynthetic proteins followed a pattern very similar to that of fatty acid synthesis proteins, implicating a role in CO(2) refixation and the supply of cofactors for oil synthesis. Expression profiles of key carbon transporters and glycolytic enzymes reflected shifts in flux from cytosolic to plastid metabolism. Despite major changes in metabolism between wri1 and wild-type seeds, <1% of genes differed by more than twofold, and most of these were involved in central lipid and carbohydrate metabolism. Thus, these data define in part the downstream responses to disruption of the WRI1 gene.

Arabidopsis↗

PubMed1998

BIOSYNTHESIS AND FUNCTION OF THE SULFOLIPID SULFOQUINOVOSYL DIACYLGLYCEROL.

The sulfolipid sulfoquinovosyl diacylglycerol is an abundant sulfur-containing nonphosphorous glycerolipid that is specifically associated with photosynthetic membranes of higher plants, mosses, ferns, algae, and most photosynthetic bacteria. The characteristic structural feature of sulfoquinovosyl diacylglycerol is the unique head group constituent sulfoquinovose, a derivative of glucose in which the 6-hydroxyl is replaced by a sulfonate group. While there is growing evidence for the final assembly of the sulfolipid by the transfer of the sulfoquinovosyl moiety from UDP-sulfoquinovose to the sn-3 position of diacylglycerol, very little is known about the biosynthesis of the precursor UDP-sulfoquinovose. Recently, a number of mutants deficient in sulfolipid biosynthesis and the corresponding sqd genes have become available from different organisms. These provide novel tools to analyze sulfolipid biosynthesis by a combination of molecular and biochemical approaches. Furthermore, the analysis of sulfolipid-deficient mutants has provided novel insights into the function of sulfoquinovosyl diacylglycerol in photosynthetic membranes.

Journal Article↗