Gene expression analysis in microdissected shoot meristems of Brassica napus microspore-derived embryos with altered SHOOTMERISTEMLESS levels

Altered expression of Brassica napus (Bn) SHOOTMERISTEMLESS (STM) affects the morphology and behaviour of microspore-derived embryos (MDEs). While down-regulation of BnSTM repressed the formation of the shoot meristem (SAM) and reduced the number of Brassica MDEs able to regenerate viable plants at germination, over-expression of BnSTM enhanced the structure of the SAM and improved regeneration frequency. Within dissected SAMs, the induction of BnSTM up-regulated the expression of many transcription factors (TFs) some of which directly involved in the formation of the meristem, i.e. CUP-SHAPED COTYLEDON1 and WUSCHEL, and regulatory components of the antioxidant response, hormone signalling, and cell wall synthesis and modification. Opposite expression patterns for some of these genes were observed in the SAMs of MDEs down-regulating BnSTM. Altered expression of BnSTM affected transcription of cell wall and lignin biosynthetic genes. The expression of PHENYLALANINE AMMONIA LYASE2, CINNAMATE 4-4HYDROXYLASE, and CINNAMYL ALCOHOL DEHYDROGENASE were repressed in SAMs over-expressing BnSTM. Since lignin formation is a feature of irreversible cell differentiation, these results suggest that one way in which BnSTM promotes indeterminate cell fate may be by preventing the expression of components of biochemical pathways involved in the accumulation of lignin in the meristematic cells. Overall, these studies provide evidence for a novel function of BnSTM in enhancing the quality of in vitro produced meristems, and propose that this gene can be used as a potential target to improve regeneration of cultured embryos.