Mutation of a mitochondrial outer membrane protein affects chloroplast lipid biosynthesis

Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI 48824, USA.
The Plant Journal (Impact Factor: 5.97). 05/2008; 54(1):163-75. DOI: 10.1111/j.1365-313X.2008.03417.x
Source: PubMed

ABSTRACT Lipid biosynthesis in plant cells is associated with various organelles, and maintenance of cell lipid homeostasis requires nimble regulation and coordination. In plants, environmental cues such as phosphate limitation require readjustment of the lipid biosynthetic machinery to substitute phospholipids by non-phosphorous glycolipids. Biosynthesis of the galactoglycerolipids predominant in plants proceeds by a constitutive and an alternative pathway that is known to be induced in response to phosphate deprivation. Plant lipid galactosyltransferases involved in both pathways are associated with the plastid envelope membranes and are encoded by nuclear genes. To identify mechanisms governing the activity of the alternative galactoglycerolipid pathway, a genetic suppressor screen was conducted in the background of the digalactolipid-deficient dgd1 mutant of Arabidopsis. A suppressor line that partially restored digalactoglycerolipid content in the dgd1 background carries a point mutation in a mitochondrial protein, which was tentatively designated DGD1 SUPPRESSOR 1 (DGS1). Presumed orthologs of this protein are present in plants, algae and fungi, but its molecular function is not yet known. In the dgd1 dgs1 double mutant, expression of nuclear genes encoding enzymes of the alternative galactoglycerolipid pathway is increased and hydrogen peroxide levels are elevated. This increase in hydrogen peroxide is proposed to be the reason for activation of the alternative pathway in the dgd1 dgs1 double mutant. Accordingly, hydrogen peroxide and treatments producing reactive oxygen also activate the alternative pathway in the wild-type. These results likely implicate the production of reactive oxygen in the regulation of the alternative galactoglycerolipid pathway in plants.

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    • "Immunoblotting was performed on polyvinylidene difluoride membranes using the CPS-1 chemiluminescent peroxidase substrate detection system from Sigma. DGS1-specific polyclonal antibodies were affinity purified as previously described (Xu et al., 2008). Monoclonal antibodies against a conserved epitope in plant AOXs (AOX mAb; Finnegan et al., 1999), and the cytochrome C oxidase subunit II (COX II mAb), were obtained from a Michigan State University-Department of Energy Plant Research Laboratory stock left by Lee McIntosh. "
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    ABSTRACT: Galactoglycerolipids are major constituents of photosynthetic membranes in chloroplasts. At least three parallel sets of enzymes are involved in their biosynthesis that must be coordinated in response to changing growth conditions. A potential candidate for a protein affecting the activity of different galactoglycerolipid pathways is the recently described digalactosyldiacylglycerol1 (dgd1) SUPPRESSOR1 (DGS1) protein of Arabidopsis (Arabidopsis thaliana) localized in the mitochondrial outer membrane. It was discovered based on a specific gain-of-function point mutation allele, dgs1-1, that causes a partial restoration of chloroplast galactoglycerolipid deficiency in the dgd1 mutant, which is defective in the lipid galactosyltransferase, DGD1. The dgs1-1 allele causes the accumulation of hydrogen peroxide that leads to an activation of an alternative, DGD1-independent galactoglycerolipid biosynthesis pathway in chloroplasts. Analysis presented here shows that the DGS1 protein is a component of a large protein complex, which explains the previously observed dominant negative phenotype following the expression of the dgs1-1 allele. The dgs1-1 allele causes the loss of mitochondrial alternative oxidase (AOX) protein that might be related to the accumulation of hydrogen peroxide in the dgs1-1 mutant background. This effect was posttranscriptional because mRNA levels for the major form of AOX were not affected in dgs1-1 mutant seedlings. Unlike dgs1-1, a loss-of-function allele, dgs1-2, had no effect on plant growth, AOX, and lipid composition to the extent tested, leaving the quest for a possible molecular function of DGS1 open. Apparently, the DGS1 wild-type protein does not directly affect lipid metabolism in mitochondria or chloroplasts.
    Plant physiology 02/2010; 152(4):1951-9. DOI:10.1104/pp.110.153262 · 6.84 Impact Factor
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    • "To construct the TGD4-GFP expression vector, the full-length coding sequence of TGD4 was amplified by RT-PCR from first-strand cDNA made from wild-type seedling mRNA, using the primers 59-CATGGATC- CATGAACAGAATGAGATGGGT-39 and 59-CACAGTCGACCTAGTGCT- CAAAGAAACGAAGC-39. Total RNA isolation and the first-strand cDNA synthesis were done as described (Xu et al., 2008). The PCR product was restricted with BamHI and SaII and inserted into the respective sites of a binary vector derived from pPZP211 carrying the GFP open reading frame, thereby creating the TGDc-GFP fusion (Hajdukiewicz et al., 1994). "
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    ABSTRACT: The development of chloroplasts in Arabidopsis thaliana requires extensive lipid trafficking between the endoplasmic reticulum (ER) and the plastid. The biosynthetic enzymes for the final steps of chloroplast lipid assembly are associated with the plastid envelope membranes. For example, during biosynthesis of the galactoglycerolipids predominant in photosynthetic membranes, galactosyltransferases associated with these membranes transfer galactosyl residues from UDP-Gal to diacylglycerol. In Arabidopsis, diacylglycerol can be derived from the ER or the plastid. Here, we describe a mutant of Arabidopsis, trigalactosyldiacylglycerol4 (tgd4), in which ER-derived diacylglycerol is not available for galactoglycerolipid biosynthesis. This mutant accumulates diagnostic oligogalactoglycerolipids, hence its name, and triacylglycerol in its tissues. The TGD4 gene encodes a protein that appears to be associated with the ER membranes. Mutant ER microsomes show a decreased transfer of lipids to isolated plastids consistent with in vivo labeling data, indicating a disruption of ER-to-plastid lipid transfer. The complex lipid phenotype of the mutant is similar to that of the tgd1,2,3 mutants disrupted in components of a lipid transporter of the inner plastid envelope membrane. However, unlike the TGD1,2,3 complex, which is proposed to transfer phosphatidic acid through the inner envelope membrane, TGD4 appears to be part of the machinery mediating lipid transfer between the ER and the outer plastid envelope membrane. The extent of direct ER-to-plastid envelope contact sites is not altered in the tgd4 mutant. However, this does not preclude a possible function of TGD4 in those contact sites as a conduit for lipid transfer between the ER and the plastid.
    The Plant Cell 09/2008; 20(8):2190-204. DOI:10.1105/tpc.108.061176 · 9.34 Impact Factor
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    ABSTRACT: Chloroplasts are the defining plant organelle carrying out photosynthesis. Photosynthetic complexes are embedded into the thylakoid membrane which forms an intricate system of membrane lamellae and cisternae. The chloroplast boundary consists of two envelope membranes controlling the exchange of metabolites between the plastid and the extraplastidic compartments of the cell. The plastid internal matrix (stroma) is the primary location for fatty acid biosynthesis in plants. Fatty acids can be assembled into glycerolipids at the envelope membranes of plastids or they can be exported and assembled into lipids at the endoplasmic reticulum (ER) to provide building blocks for extraplastidic membranes. Some of these glycerolipids, assembled at the ER, return to the plastid where they are remodeled into the plastid typical glycerolipids. As a result of this cooperation of different subcellular membrane systems, a rich complement of lipid trafficking phenomena contributes to the biogenesis of chloroplasts. Considerable progress has been made in recent years towards a better mechanistic understanding of lipid transport across plastid envelopes. Lipid transporters of bacteria and plants have been discovered and their study begins to provide detailed mechanistic insights into lipid trafficking phenomena relevant to chloroplast biogenesis.
    Progress in Lipid Research 10/2008; 47(5):381-9. DOI:10.1016/j.plipres.2008.04.001 · 10.02 Impact Factor
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