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Published on January 26, 2007; 10.1104/pp.106.090811


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Received October 3, 2006
Accepted January 3, 2007

Characterization of a Plastid Triacylglycerol Lipase from Arabidopsis

Anita K. Padham , Marianne T. Hopkins , Tzann-Wei Wang , Linda M. McNamara , Maisie Lo , Lynn G.L. Richardson , Matthew D. Smith , Catherine A. Taylor , and John E. Thompson *

Department of Biology, University of Waterloo, Waterloo, Ontario, Canada N2L 3G1; Department of Biology, Wilfrid Laurier University, Waterloo, Ontario, Canada N2L 3C5

* Corresponding author; email: jet{at}sciborg.uwaterloo.ca.

Full-length cDNA corresponding to Arabidopsis thaliana gene At2g31690, which has been annotated in GenBank as a putative triacylglycerol lipase, was obtained by RT-PCR using RNA from senescing rosette leaves of Arabidopsis as a template. The cognate protein was found to contain the lipase active site sequence, and corresponding recombinant protein proved capable of de-esterifying triacylglycerol. In vitro chloroplast import assays indicated that the lipase is targeted to chloroplasts. This was confirmed by confocal microscopy of rosette leaf tissue treated with FITC-labeled, lipase-specific antibody, which revealed that the lipase protein co-localizes with plastoglobular neutral lipid. Western blot analysis indicated that the lipase is expressed in roots, inflorescence stems, flowers, siliques and leaves, and that it is strongly up-regulated in senescing rosette leaf tissue. Transgenic plants with suppressed lipase protein levels were obtained by expressing At2g31690 cDNA in antisense orientation under the regulation of a constitutive promoter. The transgenic plants bolted and flowered at the same time as wild-type plants, but were severely stunted and exhibited delayed rosette senescence. Moreover, the stunted growth phenotype correlated with irregular chloroplast morphology. The chloroplasts of transgenic plants were structurally deformed, had a reduced abundance of thylakoids that were abnormally stacked, and contained more plastoglobular neutral lipid than chloroplasts of wild-type plants. These observations collectively indicate that this triacylglycerol lipase plays a role in maintaining the structural integrity of chloroplasts, possibly by mobilizing the fatty acids of plastoglobular triacylglycerol.







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