SummaryThe importance of the arginyl-tRNA protein transferase (ATE), the enzyme mediating posttranslation arginylation of proteins in the N-end rule degradation (NERD) pathway of protein stability, was analysed in Physcomitrella patens and compared to its known functions in other eukaryotes.We characterize ATE:GUS reporter lines as well as ATE mutants in P. patens to study the impact and function of arginylation on moss development and physiology.ATE protein abundance is spatially and temporally regulated in P. patens by hormones and light and is highly abundant in meristematic cells. Further, the amount of ATE transcript is regulated during abscisic acid signalling and downstream of auxin signalling. Loss-of-function mutants exhibit defects at various levels, most severely in developing gametophores, in chloroplast starch accumulation and senescence. Thus, arginylation is necessary for moss gametophyte development, in contrast to the situation in flowering plants.Our analysis further substantiates the conservation of the N-end rule pathway components in land plants and highlights lineage-specific features. We introduce moss as a model system to characterize the role of the NERD pathway as an additional layer of complexity in eukaryotic development.