Peroxisomal Acyl-CoA oxidase 4 activity differs between Arabidopsis accessions
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In plants, peroxisomes are the primary site of fatty acid β-oxidation. Following substrate activation, fatty acids are oxidized by Acyl-CoA Oxidase (ACX) enzymes. Arabidopsis has six ACX genes, although ACX6 is not expressed. Biochemical characterization has revealed that each ACX enzyme acts on specific chain-length targets, but in a partially overlapping manner, indicating a degree of functional redundancy. Genetic analysis of acx single and double mutants in the Columbia (Col-0) accession revealed only minor phenotypes, but an acx3acx4 double mutant from Wassileskija (Ws) is embryo lethal. In this study, we show that acx3acx4 Col and acx1acx3acx4 Col mutants are viable and that enzyme activity in these mutants is significantly reduced on a range of substrates compared to wild type. However, the triple mutant displays only minor defects in seed-storage mobilization, seedling development, and adult growth. Although the triple mutant is defective in the three most active and highly-expressed ACX proteins, increases in ACX2 expression may support partial β-oxidation activity. Comparison of acx mutant alleles in the Col-0 and Ws accessions reveals independent phenotypes; the Ws acx4 mutant uniquely shows increased sensitivity to propionate, whereas the Col-0 acx4 allele has sucrose-dependent growth in the light. To dissect the issues between Col-0 and Ws, we generated mixed background mutants. Although alleles with the Col-0 acx4 mutant were viable, we were unable to isolate an acx3acx4 line using the Ws acx4 allele. Reducing ACX4 expression in several Arabidopsis backgrounds showed a split response, suggesting that the ACX4 gene and/or protein functions differently in Arabidopsis accessions.
KeywordsAcyl-CoA oxidase ACX Peroxisome Fatty acid beta-oxidation
This research was supported by the National Science Foundation (IOS-0845507), a University of Missouri-St. Louis Research Award, and University of Missouri-St. Louis start-up funds. We are grateful to Ian Graham for supplying acx mutants in the Ws background for comparison and the Arabidopsis Biological Resource Center for the ACX4 clone. We gratefully acknowledge the assistance of Amanda Tawfall and Xuemin Wang in the performance of the GC analysis and Anthony Fischer, Melanie Miller, Wendy Olivas, and Joe Russo for assistance with northern blots. We also thank Robert Barlow for assistance in optimizing the ACX enzyme assays, Anupama Vijayaraghavan, Amanda Tulin, and Tad Wood for assistance in genotyping the acx double and triple mutants, and Steven Jarvis for help isolating RNA interference lines. We are grateful to Robert Barlow, Shelly Boyer, Ying Li, Lisa Schechter, Gretchen Spiess, Amanda Tulin, and Anupama Vijayaraghavan for critical comments on the manuscript.
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