Characterization Of An Arabidopsis Long Chain Fatty Acyl-Coenzyme A Synthetase, Which Is Required For Seedling Establishment Without Exogenous Sugar

The degradation of storage lipids is essential for seedling establishment in oilseed plants,provides energy and molecular necessities for the building of photosynthetic apparatus andallowing plant autotrophic growth. In mature stage, degradation fatty acid can remove thetoxic free fatty acids produced in the cell senescence process to ensure plant normaldevelopment. To enter the β-oxidation pathway, fatty acids need to be activated to form acylCoA catalyzed by acyl CoA synthetase. Here we report the characterization of anArabidopsis cDNA which encodes a long chain acyl-CoA synthetase (LACS, EC6.2.1.3)designated as LACS1. LACS1 consists of 660 amino acids with a calculated molecular massof 74597. Cluster analysis show that LACS1 fell into an independent group among the ninemembers in LACS family of Arabidopsis. Transgenic plants with suppressed LACS1expression could not carry out the seedling establishment without the supplement ofexogenous sugars and the lipid degradation during germination was inhibited, fatty acid assyrevealed that that LACS1 has the preference to catalyze the saturation fatty acids. Overexpression of LACS1 in Arabidopsis leads to the increase of the seed lipids content, whilesuppression of LACS1 results in the decrease of the lipid content, but the composition of fattyacids in two transgenic plant seeds was not altered. The LACS1 involved in fatty acidbreakdown, but did not involved in peroxisomal β-oxidation, in addition, it has amitochondrial signal in N terminus, and LACS1-GFP fusion protein transiently expressed inthe cytoplasm in the onion epidermal cell, deducing that it may act in mitochondrialβ-oxidation pathway. PT-PCR and Northern analysis show that LACS1 expressed increase to ahigh level at 3 days to 5 days after germination and then decrease in adult plants, LACS1Abstract 3mainly expressed in flowers, inflorescence and stems. These data revealed that LACS1 playan important role in fatty acid degradation and seedling establishment, also it has an effects onadult plant development. In order to utilize the At-LACS1 in oil rape to increase the oil content, we use partialsequence of At-LACS1 as a probe to hybridize the TAC genomic library of Brassica napus, 35positive clones were gotten. The positive clones fell into 5 types by Southern blotting analysis,revealed that Brassica napus genome contains 5 homologue genes of At-LACS1. The functionof seed specific promoter Napin cloned by our lab was confirmed in Arabidopsis, it can drivegene expression in the seed derived cotyledons and hypocotyl in seedling stage, and in theskins of siliques in the mature stage.