Schnurr Judy, Shockey Jay, Browse John
Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164, USA.
Plant Cell. 2004 Mar;16(3):629-42. doi: 10.1105/tpc.017608. Epub 2004 Feb 18.
Long-chain acyl-CoA synthetase (LACS) activities are encoded by a family of at least nine genes in Arabidopsis (Arabidopsis thaliana). These enzymes have roles in lipid synthesis, fatty acid catabolism, and the transport of fatty acids between subcellular compartments. Here, we show that the LACS2 gene (At1g49430) is expressed in young, rapidly expanding tissues, and in leaves expression is limited to cells of the adaxial and abaxial epidermal layers, suggesting that the LACS2 enzyme may act in the synthesis of cutin or cuticular waxes. A lacs2 null mutant was isolated by reverse genetics. Leaves of mutant plants supported pollen germination and released chlorophyll faster than wild-type leaves when immersed in 80% ethanol, indicating a defect in the cuticular barrier. The composition of surface waxes extracted from lacs2 leaves was similar to the wild type, and the total wax load was higher than the wild type (111.4 microg/dm(2) versus 76.4 microg/dm(2), respectively). However, the thickness of the cutin layer on the abaxial surface of lacs2 leaves was only 22.3 +/- 1.7 nm compared with 33.0 +/- 2.0 nm for the wild type. In vitro assays showed that 16-hydroxypalmitate was an excellent substrate for recombinant LACS2 enzyme. We conclude that the LACS2 isozyme catalyzes the synthesis of omega-hydroxy fatty acyl-CoA intermediates in the pathway to cutin synthesis. The lacs2 phenotype, like the phenotypes of some other cutin mutants, is very pleiotropic, causing reduced leaf size and plant growth, reduced seed production, and lower rates of seedling germination and establishment. The LACS2 gene and the corresponding lacs2 mutant will help in future studies of the cutin synthesis pathway and in understanding the consequences of reduced cutin production on many aspects of plant biology.
长链脂酰辅酶A合成酶（LACS）活性由拟南芥（Arabidopsis thaliana）中至少9个基因组成的家族编码。这些酶在脂质合成、脂肪酸分解代谢以及亚细胞区室之间的脂肪酸转运中发挥作用。在此，我们表明LACS2基因（At1g49430）在幼嫩、快速生长的组织中表达，在叶片中表达仅限于近轴和远轴表皮层的细胞，这表明LACS2酶可能在角质或表皮蜡质的合成中起作用。通过反向遗传学分离出一个lacs2缺失突变体。当浸入80%乙醇中时，突变体植株的叶片比野生型叶片更能支持花粉萌发且叶绿素释放更快，表明角质层屏障存在缺陷。从lacs2叶片中提取的表面蜡质成分与野生型相似，且总蜡质负载高于野生型（分别为111.4微克/平方分米和76.4微克/平方分米）。然而，lacs2叶片远轴表面角质层的厚度仅为22.3±1.7纳米，而野生型为33.0±2.0纳米。体外试验表明，16-羟基棕榈酸是重组LACS2酶的优良底物。我们得出结论，LACS2同工酶在角质合成途径中催化ω-羟基脂肪酰辅酶A中间体的合成。lacs2表型与其他一些角质突变体的表型一样，具有很强的多效性，导致叶片大小和植株生长减小、种子产量降低以及幼苗萌发和定植率降低。LACS2基因和相应的lacs2突变体将有助于未来对角质合成途径的研究，并有助于理解角质产量降低对植物生物学许多方面的影响。
