Membrane-bound glycerol-3-phosphate acyltransferase (GPAT; EC 2.3.1.15) mediates the initial step of glycerolipid biosynthesis in the extraplastidic compartments of plant cells. Here, we report the molecular characterization of a novel GPAT gene family from Arabidopsis, designated AtGPAT. The corresponding polypeptides possess transmembrane domains and GPAT activity when expressed heterologously in a yeast lipid mutant. The functional significance of one isoform, AtGPAT1, is the focus of the present study. Disruption of the AtGPAT1 gene causes a massive pollen development arrest, and subsequent introduction of the gene into the mutant plant rescues the phenotype, illustrating a pivotal role for AtGPAT1 in pollen development. Microscopic examinations revealed that the gene lesion results in a perturbed degeneration of the tapetum, which is associated with altered endoplasmic reticulum profiles and reduced secretion. In addition to the sporophytic effect, AtGPAT1 also exerts a gametophytic effect on pollen performance, as the competitive ability of a pollen grain to pollinate is dependent on the presence of an AtGPAT1 gene. Deficiency in AtGPAT1 correlates with several fatty acid composition changes in flower tissues and seeds. Unexpectedly, however, a loss of AtGPAT1 causes no significant change in seed oil content.

In higher plants, three subfamilies of sucrose nonfermenting-1 (Snf1)-related protein kinases have evolved. While the Snf1-related protein kinase 1 (SnRK1) subfamily has been shown to share pivotal roles with the orthologous yeast Snf1 and mammalian AMP-activated protein kinase in modulating energy and metabolic homeostasis, the functional significance of the two plant-specific subfamilies SnRK2 and SnRK3 in these critical processes is poorly understood. We show here that SnRK2.6, previously identified as crucial in the control of stomatal aperture by abscisic acid (ABA), has a broad expression pattern and participates in the regulation of plant primary metabolism. Inactivation of this gene reduced oil synthesis in Arabidopsis (Arabidopsis thaliana) seeds, whereas its overexpression increased Suc synthesis and fatty acid desaturation in the leaves. Notably, the metabolic alterations in the SnRK2.6 overexpressors were accompanied by amelioration of those physiological processes that require high levels of carbon and energy input, such as plant growth and seed production. However, the mechanisms underlying these functionalities could not be solely attributed to the role of SnRK2.6 as a positive regulator of ABA signaling, although we demonstrate that this kinase confers ABA hypersensitivity during seedling growth. Collectively, our results suggest that SnRK2.6 mediates hormonal and metabolic regulation of plant growth and development and that, besides the SnRK1 kinases, SnRK2.6 is also implicated in the regulation of metabolic homeostasis in plants.

To what extent fatty acyltransferases control fatty acid composition in glycerolipids is largely unknown. To gain some insight into this process, we characterized a membrane-bound glycerol-3-phosphate acyltransferase (GPAT), designated TpGPAT, from the marine diatom Thalassiosira pseudonana Hasle & Heim. The glycerolipids from T. pseudonana consist predominantly of 16:0, 16:1ω7, and 20:5ω3 fatty acids. Heterologous expression of TpGPAT in a yeast GPAT-deficient mutant (gat1) demonstrated that TpGPAT could effectively use glycerol-3-phosphate, but not dihydroxyacetone phosphate, as fatty acyl acceptor. This enzyme highly preferred 16:0 in an in vitro enzyme assay, and discriminated against the monounsaturated 16-carbon fatty acid. Accordingly, expression of TpGPAT in gat1 resulted in approximately 18% and 12% increases of 16:0 in triacylglycerols and phospholipids, respectively. The unsaturated fatty acids, 16:1 and 18:1, on the other hand, were reduced by 15% and 21% in these two lipid species. Collectively, the results demonstrate that TpGPAT possesses a high substrate preference for 16:0-CoA and it exerts a large effect on the fatty-acid composition of glycerolipids.