Cuticular waxes are complex mixtures of very-long-chain fatty acids (VLCFAs) and their derivatives, forming a natural barrier on aerial surfaces of terrestrial plants against biotic and abiotic stresses. In VLCFA biosynthesis, β-ketoacyl-CoA synthase (KCS) is the key enzyme, catalyzing the first reaction in fatty acid elongation and determining substrate specificity. We isolated a rice (Oryza sativa) wax crystal-sparse leaf 4 (WSL4) gene using a map-based cloning strategy. WSL4 is predicted to encode a KCS, a homolog of Arabidopsis (Arabidopsis thaliana) CER6. Complementation of the mutant wsl4-1 with WSL4 genomic DNA rescued the cuticular wax-deficient phenotype, confirming the function of WSL4 The load of wax components longer than 30 carbons (C30) and C28 were reduced markedly in wsl4-1 and wsl4-2 mutants, respectively. Overexpression of WSL4 increased the cuticular wax load in rice leaves. We further isolated a cofactor of WSL4, OsCER2, a homolog of Arabidopsis CER2, by coimmunoprecipitation and confirmed their physical interaction by split-ubiquitin yeast two-hybrid experiments. Expression of WSL4 alone in elo3 yeast cells resulted in increased C24 but did not produce VLCFAs of greater length, whereas expressing OsCER2 alone showed no effect. Coexpression of WSL4 and OsCER2 in elo3 yeast cells yielded fatty acids up to C30. OsCER2 with a mutated HxxxD motif (H172E, D176A, and D176H) interrupted its interaction with WSL4 and failed to elongate VLCFAs past C24 when expressed with WSL4 in elo3 yeast cells. These results demonstrated that WSL4 was involved in VLCFA elongation beyond C22 and that elongation beyond C24 required the participation of OsCER2.

Here we report two unrelated Chinese families with congenital missing teeth inherited in an X-linked manner. We mapped the affected locus to chromosome Xp11-Xq21 in one family. In the defined region, both families were found to have novel missense mutations in the ectodysplasin-A (EDA) gene. The mutation of c.947A>G caused the D316G substitution of the EDA protein. The mutation of c.1013C>T found in the other family resulted in the Thr to Met mutation at position 338 of EDA. The EDA gene has been reported responsible for X-linked hypohidrotic ectodermal dysplasia (XLHED) in humans characterized by impaired development of hair, eccrine sweat glands, and teeth. In contrast, all the affected individuals in the two families that we studied here had normal hair and skin. Structural analysis suggests that these two novel mutants may account for the milder phenotype by affecting the stability of EDA trimers. Our results indicate that these novel missense mutations in EDA are associated with the isolated tooth agenesis and provide preliminary explanation for the abnormal clinical phenotype at a molecular structural level.

Plastidial disproportionating enzyme1 (DPE1), an a-1,4-D-glucanotransferase, has been thought to be involved in storage starch synthesis in cereal crops. However, the precise function of DPE1 remains to be established. We present here the functional identification of DPE1 in storage starch synthesis in rice (Oryza sativa) by endosperm-specific gene overexpression and suppression. DPE1 overexpression decreased amylose content and resulted in small and tightly packed starch granules, whereas DPE1 suppression increased amylose content and formed heterogeneous-sized, spherical, and loosely packed starch granules. Chains with degree of polymerization (DP) of 6 to 10 and 23 to 38 were increased, while chains with DP of 11 to 22 were decreased in amylopectin from DPE1-overexpressing seeds. By contrast, chains with DP of 6 to 8 and 16 to 36 were decreased, while chains with DP of 9 to 15 were increased in amylopectin from DPE1-suppressed seeds. Changes in DPE1 gene expression also resulted in modifications in the thermal and pasting features of endosperm starch granules. In vitro analyses revealed that recombinant DPE1 can break down amylose into maltooligosaccharides in the presence of Glc, while it can transfer maltooligosyl groups from maltooligosaccharide to amylopectin or transfer maltooligosyl groups within and among amylopectin molecules in the absence of Glc. Moreover, a metabolic flow of maltooligosyl groups from amylose to amylopectin was clearly identifiable when comparing DPE1-overexpressing lines with DPE1-suppressed lines. These findings demonstrate that DPE1 participates substantially in starch synthesis in rice endosperm by transferring maltooligosyl groups from amylose and amylopectin to amylopectin.Starch is important for plant development and is critical for crop quality and nutrition. The starch accumulated in cereal endosperm is the most important dietary source of energy for humans. Starch consists of two types of D-Glc homopolymers, amylose and amylopectin, accounting for 15% to 30% and 70% to 85%, respectively (Hermansson and Svegmark, 1996;Tester et al., 2004). Amylose is a largely linear polymer, in which glucosyl monomers are joined via a-1,4-glycoside bonds. Amylopectin is a highly branched molecule with 5% to