RESUMO
CGI-58 is the defective gene in the human neutral lipid storage disease called Chanarin-Dorfman syndrome. This disorder causes intracellular lipid droplets to accumulate in nonadipose tissues, such as skin and blood cells. Here, disruption of the homologous CGI-58 gene in Arabidopsis thaliana resulted in the accumulation of neutral lipid droplets in mature leaves. Mass spectroscopy of isolated lipid droplets from cgi-58 loss-of-function mutants showed they contain triacylglycerols with common leaf-specific fatty acids. Leaves of mature cgi-58 plants exhibited a marked increase in absolute triacylglycerol levels, more than 10-fold higher than in wild-type plants. Lipid levels in the oil-storing seeds of cgi-58 loss-of-function plants were unchanged, and unlike mutations in ß-oxidation, the cgi-58 seeds germinated and grew normally, requiring no rescue with sucrose. We conclude that the participation of CGI-58 in neutral lipid homeostasis of nonfat-storing tissues is similar, although not identical, between plant and animal species. This unique insight may have implications for designing a new generation of technologies that enhance the neutral lipid content and composition of crop plants.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Lipídeos/química , Folhas de Planta/metabolismo , 1-Acilglicerol-3-Fosfato O-Aciltransferase/genética , 1-Acilglicerol-3-Fosfato O-Aciltransferase/metabolismo , Fatores Etários , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Primers do DNA/genética , Ácidos Graxos/análise , Humanos , Espectrometria de Massas , Microscopia de Fluorescência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sementes/química , Triglicerídeos/análiseRESUMO
An expanding appreciation for the varied functions of neutral lipids in cellular organisms relies on a more detailed understanding of the mechanisms of lipid production and packaging into cytosolic lipid droplets (LDs). Conventional lipid profiling procedures involve the analysis of tissue extracts and consequently lack cellular or subcellular resolution. Here, we report an approach that combines the visualization of individual LDs, microphase extraction of lipid components from droplets, and the direct identification of lipid composition by nanospray mass spectrometry, even to the level of a single LD. The triacylglycerol (TAG) composition of LDs from several plant sources (mature cotton (Gossypium hirsutum) embryos, roots of cotton seedlings, and Arabidopsis thaliana seeds and leaves) were examined by direct organelle mass spectrometry and revealed the heterogeneity of LDs derived from different plant tissue sources. The analysis of individual LDs makes possible organellar resolution of molecular compositions and will facilitate new studies of LD biogenesis and functions, especially in combination with analysis of morphological and metabolic mutants. Furthermore, direct organelle mass spectrometry could be applied to the molecular analysis of other subcellular compartments and macromolecules.