ECERIFERUM2-LIKE proteins have unique biochemical and physiological functions in very-long-chain fatty acid elongation.

Tegan M Haslam,Jérôme Joubès,Johnathan A Napier,Camille Delude,Frédéric Beaudoin,Ljerka Kunst,Didier Thoraval,Frédéric Domergue,Aurora Mañas Fernández,Richard Haslam,Stéphanie Pascal

doi:10.1104/pp.114.253195

Abstract

The extension of very-long-chain fatty acids (VLCFAs) for the synthesis of specialized apoplastic lipids requires unique biochemical machinery. Condensing enzymes catalyze the first reaction in fatty acid elongation and determine the chain length of fatty acids accepted and produced by the fatty acid elongation complex. Although necessary for the elongation of all VLCFAs, known condensing enzymes cannot efficiently synthesize VLCFAs longer than 28 carbons, despite the prevalence of C28 to C34 acyl lipids in cuticular wax and the pollen coat. The eceriferum2 (cer2) mutant of Arabidopsis (Arabidopsis thaliana) was previously shown to have a specific deficiency in cuticular waxes longer than 28 carbons, and heterologous expression of CER2 in yeast (Saccharomyces cerevisiae) demonstrated that it can modify the acyl chain length produced by a condensing enzyme from 28 to 30 carbon atoms. Here, we report the physiological functions and biochemical specificities of the CER2 homologs CER2-LIKE1 and CER2-LIKE2 by mutant analysis and heterologous expression in yeast. We demonstrate that all three CER2-LIKEs function with the same small subset of condensing enzymes, and that they have different effects on the substrate specificity of the same condensing enzyme. Finally, we show that the changes in acyl chain length caused by each CER2-LIKE protein are of substantial importance for cuticle formation and pollen coat function.

Highlights

The extension of very-long-chain fatty acids (VLCFAs) for the synthesis of specialized apoplastic lipids requires unique biochemical machinery
Coexpression of CER2, CER2-LIKE1, and CER6, or CER2-LIKE1, CER2-LIKE2, and CER6, produced VLCFA profiles similar to CER2-LIKE1 and CER6 alone, indicating that there is no synergistic effect of having multiple CER2-LIKE proteins
These results demonstrate that CER2-LIKE proteins have unique substrate specificities in the presence of the same condensing enzyme

Summary

Introduction

The extension of very-long-chain fatty acids (VLCFAs) for the synthesis of specialized apoplastic lipids requires unique biochemical machinery. The eceriferum (cer2) mutant of Arabidopsis (Arabidopsis thaliana) was previously shown to have a specific deficiency in cuticular waxes longer than 28 carbons, and heterologous expression of CER2 in yeast (Saccharomyces cerevisiae) demonstrated that it can modify the acyl chain length produced by a condensing enzyme from 28 to 30 carbon atoms. Eceriferum (cer2) and glossy (gl2) mutants of Arabidopsis and Zea mays, respectively, are deficient in specific VLCFA-derived waxes longer than C28 (Bianchi et al, 1975; McNevin et al, 1993; Jenks et al, 1995) Both mutations were mapped to genes that do not resemble any component of the elongase complex (Tacke et al, 1995; Xia et al, 1996), but are homologous to the BAHD family of acyltransferases (St-Pierre et al, 1998). Site-directed mutagenesis of conserved acyltransferase catalytic site amino acids in CER2 revealed that this motif is not required for CER2 function in cuticular wax synthesis (Haslam et al, 2012)

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Conclusion