One of the most exciting discoveries in recent years, as regards cell wall formation, is that of the roles of two transcription factors [SND1 (also called NST3) and NST1]. These are responsible for (secondary wall formation) in fibers of Arabidopsis (198, 199), and are specifically expressed in interfascicular fibers and xylary fibers as shown using the GUS reporter gene strategy (198,199). Analysis of the SND1 (NST3) knockout line, however, did not initially indicate any anatomical differences when compared to the wild type, this being attributed to possible genetic redundancy. To overcome this problem, a dominant repression strategy (198) was next used where SND1 (NST3) was fused to the EAR repression domain (283). Fifteen of the 64 transgenic plants obtained displayed a phenotype that was unable to stand upright. Cross-sections of the stems showed that the interfascicular fibers and xylary fibers were very thin, due to a lack of secondary wall development; the cell walls of the vessels by comparison were unaffected.

Analysis of SND1 (NST3) and NST1 promoter activity also indicated that both transcrip­tion features were expressed in fibers suggesting that both may be involved in secondary wall thickening (199). A double mutant, nst1-1 nst3-1, was next generated, with the result­ing phenotypes such that the plants obtained were unable to remain upright after reaching ~15 cm in length (Figure 7.13F). Stem cross-sections also indicated that the interfascicular fibers were not autofluorescent (an indication of lignification) compared to wild type, and ultrastructural analyses established the absence of secondary cell walls in the fibers; again vessels were unaffected as regards normal secondarycell wall development. Quantitative real time PCR analyses also indicated that various genes involved in secondary wall biosynthesis were suppressed (198,199), these included those involved in monolignol (CCOMT, At4CL1, AtOMTl, AtCCR1), as well as cellulose (irx3, irx5) and xylan (fra8) biosynthesis. This is a very important discovery, not only that fiber secondary wall formation is under control of transcriptional factors, but also that the plant lines are again unable to compensate in any effective manner for the defects introduced.