In situ analysis of lignins in transgenic tobacco reveals a differential impact of individual transformations on the spatial patterns of lignin deposition at the cellular and subcellular levels

Using tobacco transgenic lines altered in the monolignol biosynthetic pathw ay and which differ in their lignin profiles we have evaluated lignin depos ition at the cellular and subcellular levels using several microanalytical techniques, Surprisingly, whereas a Cinnamoyl CoA reductase (CCR) down-regu lated line with a strong decrease in lignin content exhibited an overall re duction in lignin deposition in the walls of the different xylem cell types , this reduction was selectively targeted to the fibers in a double transfo rmant (down-regulated for both CCR and Cinnamyl alcohol dehydrogenase (CAD) ) displaying a similar degree of global lignin content decrease. Fiber and vessel secondary walls of the transgenic tobacco line homozygous for the cc r antisense gene (CCR.H) down-regulated plants were dramatically destructur ed, particularly in the S2 sublayer, whereas the deposition of lignins in t he S1 sublayer was not significantly modified. In contrast, cell wall organ ization was slightly altered in xylem cells of the double transformant. The relative distribution of non-condensed and condensed units in lignin, eval uated microscopically with specific antibodies, was differentially affected in the transgenics studied and, in a general way, a drop in non-condensed lignin units (beta- 0-4 interunit linkages) was associated with a loss of c ohesion and extensive disorganization of the secondary wall. These results demonstrate that lignification is tightly and independently regulated in in dividual cell types and cell wall sublayers. They also show that down-regul ation of specific genes may induce targeted changes in lignin structure and in spatial deposition patterns of the polymer.