Reorganization of wall components is required to allow growth and differentiation. One matrix polysaccharide that is postulated to play an important role in this reorganization is xyloglucan (XyG). While the structure of XyG is well understood, its biosynthesis is not. Through genetic studies with Arabidopsis CSLC genes, we demonstrate that they are responsible for the synthesis of the XyG glucan backbone. A quintuple cslc mutant is able to grow and develop normally but lacks detectable XyG. These results raise important questions regarding cell wall structure and its reorganization during growth. The series of cslc mutants will be valuable tools for investigating these questions.
Xyloglucan (XyG) is an abundant component of the primary cell walls of most plants. While the structure of XyG has been well studied, much remains to be learned about its biosynthesis. Here we employed reverse genetics to investigate the role of Arabidopsis cellulose synthase like-C (CSLC) proteins in XyG biosynthesis. We found that single mutants containing a T-DNA in each of the five Arabidopsis CSLC genes had normal levels of XyG. However, higher-order cslc mutants had significantly reduced XyG levels, and a mutant with disruptions in all five CSLC genes had no detectable XyG. The higher-order mutants grew with mild tissue-specific phenotypes. Despite the apparent lack of XyG, the cslc quintuple mutant did not display significant alteration of gene expression at the whole-genome level, excluding transcriptional compensation. The quintuple mutant could be complemented by each of the five CSLC genes, supporting the conclusion that each of them encodes a XyG glucan synthase. Phylogenetic analyses indicated that the CSLC genes are widespread in the plant kingdom and evolved from an ancient family. These results establish the role of the CSLC genes in XyG biosynthesis, and the mutants described here provide valuable tools with which to study both the molecular details of XyG biosynthesis and the role of XyG in plant cell wall structure and function.
Figure 1: Isolation of cslc single mutants. (A) Gene models of all five cslc single mutants. Black rectangles, exons; black lines, introns; red triangles, T-DNA insertion sites; blue lines, noncoding sequences. Numbers in the red triangles indicate alleles. (B) The genotype of each cslc mutant was confirmed by PCR using gene- and T-DNA–specific primers. LP and RP primers were used to amplify a portion of the genomic sequence of each CSLC gene. LB is the specific primer on the T-DNA used to verify the presence of T-DNA. In each panel, DNA amplification is presented for the wild type (first lane) and three biological replicates for each single mutant.