Differentiating xylem, the water-conducting tissue in plants, has long been known to undergo programmed cell death (PCD). The transformation from a living cell into a hollow water-conducting vessel involves cell elongation, formation of secondary cell walls followed by PCD through rupturing of the vacuole and digestion of the nucleus and organelles. A group of proteases called metacaspases have been suggested to play an important role in PCD in plants. Metacaspase 9 have been shown to be upregulated in Populus during xylem maturation and also apperars to be under direct control of Vascular-related NAC-domain protein 7 (VND7), a master regulator of secondary wall biosynthesis in xylem vessels. Therefore this study aimed at identifying upstream actors of metacaspase 9 and learn more about the genes affecting xylem development and PCD.

By using a reporter line based on Columbia ecotype of Arabidopsis thaliana where green fluorescent protein (GFP) was connected to the promoter of metacaspase 9, AtMC9::nGFP, the expression of this protease could be traced in living plant tissues. The reporter line was also used to create mutants through ethyl methanesulfonate seed mutagenesis searching for mutants with an altered GFP pattern as these were carriers of mutations affecting the expression of metacaspase 9. Mapping and deep sequencing was used to locate the positions of the mutations and identify candidate genes.