title: TGD4 involved in endoplasmic reticulum-to-chloroplast lipid trafficking is a phosphatidic acid binding protein.
abstract: The synthesis of galactoglycerolipids, which are prevalent in photosynthetic membranes, involves enzymes at the endoplasmic reticulum (ER) and the chloroplast envelope membranes. Genetic analysis of trigalactosyldiacylglycerol (TGD) proteins in Arabidopsis has demonstrated their role in polar lipid transfer from the ER to the chloroplast. The TGD1, 2, and 3 proteins resemble components of a bacterial-type ATP-binding cassette (ABC) transporter, with TGD1 representing the permease, TGD2 the substrate binding protein, and TGD3 the ATPase. However, the function of the TGD4 protein in this process is less clear and its location in plant cells remains to be firmly determined. The predicted C-terminal β-barrel structure of TGD4 is weakly similar to proteins of the outer cell membrane of Gram-negative bacteria. Here, we show that, like TGD2, the TGD4 protein when fused to DsRED specifically binds phosphatidic acid (PtdOH). As previously shown for tgd1 mutants, tgd4 mutants have elevated PtdOH content, probably in extraplastidic membranes. Using highly purified and specific antibodies to probe different cell fractions, we demonstrated that the TGD4 protein was present in the outer envelope membrane of chloroplasts, where it appeared to be deeply buried within the membrane except for the N-terminus, which was found to be exposed to the cytosol. It is proposed that TGD4 is either directly involved in the transfer of polar lipids, possibly PtdOH, from the ER to the outer chloroplast envelope membrane or in the transfer of PtdOH through the outer envelope membrane.
authors.affiliation: Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI 48824, USA.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Wang
authors.forename: Changcheng
authors.initials: C
authors.lastname: Xu
authors.forename: Christoph
authors.initials: C
authors.lastname: Benning
mesh_terms: D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D015151:Immunoblotting; D007425:Intracellular Membranes / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008563:Membrane Lipids / Q000378:metabolism*; D009154:Mutation; D010712:Phosphatidic Acids / Q000378:metabolism*; D030821:Plants, Genetically Modified; D011485:Protein Binding
chemical_list: D029681:Arabidopsis Proteins; D002352:Carrier Proteins; D008164:Luminescent Proteins; D008563:Membrane Lipids; D010712:Phosphatidic Acids; C534346:TGD4 protein, Arabidopsis; C413663:fluorescent protein 583