Although chloroplasts can synthesize fatty acids and PA, PA originating from the ER can also be imported from the cytosol by the trigalactosyldiacylglycerol complex. At the stromal side of the chloroplast envelope, PA is converted to diacylglycerol by PA phosphatases; diacylglycerol is then used as a substrate for MDGD synthase. In published graphical representations of the lipid transport routes, the path of PA from the ER to the chloroplast has often been represented with dashed lines or a question mark, indicating that the factors mediating the transport were unknown. Sec14 proteins, however, represented good candidates since they have well-established functions in lipid transport.
The researchers identified two Sec14 proteins, SFH5 and SFH7, that have a high expression level in cotyledons and leaves. Only double mutants are visibly smaller and paler than the control, with reduced levels of chlorophyll and an aberrant thylakoid organization, suggesting redundant functions of these two proteins in chloroplast development. It is furthermore shown that the recombinant Sec14 domain of SFH5 can bind PA and, to a lesser extent, phosphatidylinositol bis- and trisphosphates. Extraction and transport assays proved that SFH5 can extract PA from, and transfer it between, membranes. The residues that are relevant for PA binding were determined by crystal structure analysis and are conserved between SFH5 and SFH7. Both proteins localize to ER and chloroplast envelope membranes, and mutants accumulate significantly less PA and MGDG. These findings close a gap in the knowledge surrounding lipid transport in plant cells.
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