Alterations in CER6, a Gene Identical to CUT1, Differentially Affect Long-Chain Lipid Content on the Surface of Pollen and Stems

Abstract

Very long chain lipids contribute to the hydrophobic cuticle on the surface of all land plants and are an essential component of the extracellular pollen coat in the Brassicaceae. Mutations in Arabidopsis CER genes eliminate very long chain lipids from the cuticle surface and, in some cases, from the pollen coat. In Arabidopsis, the loss of pollen coat lipids can disrupt interactions with the stigma, inhibiting pollen hydration and causing sterility. We have positionally cloned CER6 and demonstrate that a wild-type copy complements the cer6-2 defect. In addition , we have identified a fertile, intragenic suppressor, cer6-2R , that partially restores pollen coat lipids but does not rescue the stem wax defect, suggesting an intriguing difference in the requirements for CER6 activity on stems and the pollen coat. Importantly, analysis of this suppressor demonstrates that low amounts of very long chain lipids are sufficient for pollen hydration and germination. The predicted CER6 amino acid sequence resembles that of fatty acid-condensing enzymes, consistent with its role in the production of epicuticular and pollen coat lipids Ͼ 28 carbons long. DNA sequence analysis revealed the nature of the cer6-1 , cer6-2 , and cer6-2R mutations, and segregation analysis showed that CER6 is identical to CUT1 , a cDNA previously mapped to a different chromosome arm. Instead, we have determined that a new gene, CER60 , with a high degree of nucleotide and amino acid similarity to CER6 , resides at the original CUT1 locus.

Cloning and Characterization of the WAX2 Gene of Arabidopsis Involved in Cuticle Membrane and Wax Production

“…Similarly, reduced fertility in cer1 and cer6 also was male gamete specific and associated with the alteration of certain pollen coat lipids (Preuss et al, 1993;Aarts et al, 1995;Fiebig et al, 2000). Evidence that altered pollen lipids disrupt normal pollen-stigma communication includes the following: (1) cer1 and cer6 pollen induces noncompatibility reactions (callose synthesis) in normally compatible stigmas; (2) wild-type, cer1, and cer6 pollen germinate in an artificial medium equally well; and (3) wild-type, cer1, and cer6 pollen placed on the same stigma induce the secretion of stigma water, leading to pollen hydration and almost normal development of the wild-type and mutant pollen.…”

“…Four of these-CER2 , CER3 , GL2 , and GL15 -appear to encode regulatory loci (Tacke et al, 1995;Hannoufa et al, 1996;Moose and Sisco, 1996;Negruk et al, 1996;Xia et al, 1996), three-CER6 , KCS1 , and GL8 -may encode metabolic enzymes (Xu et al, 1997;Millar et al, 1999;Todd et al, 1999;Fiebig et al, 2000), and two-CER1 and GL1 -may be involved in either the transport or the metabolism of wax compounds (Aarts et al, 1995;Hansen et al, 1997). None of these genes has been associated with cuticle membrane synthesis.…”

The Cytochrome P450 Enzyme CYP96A15 Is the Midchain Alkane Hydroxylase Responsible for Formation of Secondary Alcohols and Ketones in Stem Cuticular Wax of Arabidopsis

“…For example, visual screenings of mutagenized Arabidopsis populations have yielded a set of 21 eceriferum (cer) mutants with characteristic changes in the composition of inflorescence stem wax (Koornneef et al, 1989;McNevin et al, 1993). Some of the genes mutated in these cer lines have been cloned and characterized, including those coding for the ketoacyl-CoA synthase CER6 (Millar et al, 1999;Fiebig et al, 2000;Hooker et al, 2002) and the enoylCoA reductase CER10 (Zheng et al, 2005) involved in fatty acid elongation, as well as the fatty acyl-CoA reductase CER4 that is responsible for primary alcohol synthesis (Rowland et al, 2006). However, not a single step in the decarbonylation pathway has been confirmed through the identification, cloning, and characterization of the predicted enzyme.…”

Disruption of Glycosylphosphatidylinositol-Anchored Lipid Transfer Protein Gene Altered Cuticular Lipid Composition, Increased Plastoglobules, and Enhanced Susceptibility to Infection by the Fungal Pathogen Alternaria brassicicola

“…During this process, the C16 and C18 fatty acids synthesized in the plastids are exported to the cytosol, where they are further elongated to VLCFAs in the range of 20 to 34 carbons by the fatty acid elongase complex on the endoplasmic reticulum (ER; Millar and Kunst, 1997;Millar et al, 1999;Todd et al, 1999;Yephremov et al, 1999;Fiebig et al, 2000;Clemens and Kunst, 2001;Hooker et al, 2002;Dietrich et al, 2005;Zheng et al, 2005). The elongated VLCFAs are then further transformed by two principal wax biosynthetic pathways: an acyl reduction pathway that produces primary alcohols and wax esters, and a decarbonylation pathway that leads to the formation of aldehydes, alkanes, secondary alcohols, and ketones (Aarts et al, 1995;Kunst and Samuels, 2003;Rowland et al, 2006).…”