Abstract
AbstractPlant proteomes show remarkable plasticity in reaction to environmental challenges and during developmental transitions. Some of this adaptability comes from ubiquitin-mediated protein destruction regulated by cullin-RING E3 ubiquitin ligases (CRLs). CRLs are activated through modification of the cullin subunit with the ubiquitin-like protein RUB/NEDD8 by an E3 ligase called defective in cullin neddylation 1 (DCN1). Here we show that tobacco DCN1 binds ubiquitin and RUB/NEDD8, and associates with cullin. When knocked down by RNAi, tobacco pollen formation stopped and zygotic embryogenesis was blocked around the globular stage. Additionally, we found that RNAi of DCN1 inhibited the stress-triggered reprogramming of cultured microspores from their intrinsic gametophytic mode of development to an embryogenic state. This stress-induced developmental switch is a known feature in many important crops and leads ultimately to the formation of haploid embryos and plants. Compensating the RNAi effect by re-transformation with a promoter-silencing construct restored pollen development and zygotic embryogenesis, as well as the ability for stress-induced formation of embryogenic microspores. Overexpression of DCN1, however, accelerated pollen tube growth and increased the potential for microspore reprogramming. These results demonstrate that the biochemical function of DCN1 is conserved in plants and that its activity is specifically required for transitions during pollen development and embryogenesis, and for pollen tube tip growth.
Highlights
Plant reproduction is characterized by a number of important developmental transitions, beginning with the acquisition of polarity in the globular embryo [1]
Overexpression of DEFECTIVE IN CULLIN NEDDYLATION 1 (DCN1), accelerated pollen tube growth and increased the potential for microspore reprogramming. These results demonstrate that the biochemical function of DCN1 is conserved in plants and that its activity is required for transitions during pollen development and embryogenesis, and for pollen tube tip growth
A pull-down experiment using At CUL1 shows that DCN1 associates with cullin (Figure 1c and Supporting Information Fig. 1), confirming data obtained in yeast and nematodes [26], [28]
Summary
Plant reproduction is characterized by a number of important developmental transitions, beginning with the acquisition of polarity in the globular embryo [1]. Instead of developing into pollen, stressed microspores become totipotent and develop into haploid embryos and plants under non-stress conditions [7] This process corresponds to a fundamental developmental process in plants, i.e. the transition of a gametophyte into a sporophyte, but can be seen as the reprogramming of a cell with a restricted development fate to a totipotent state. Rubylation/neddylation and derubylation/deneddylation have been shown to be essential in plants, worms, and mammals, partially in fission yeast, but not in budding yeast (reviewed by [9], [21]) They play a fundamental role in important processes such as morphogenesis [22], cell division [23], signaling [24] and embryogenesis [25],[26]. Using RNAi knockdown we show that DCN1 controls the key developmental transitions from microsporogenesis to microgametogenesis, from gametophytic to sporophytic development in cultured microspores, and from the globular to the heart-shaped stage in both zygotic and microspore embryogenesis
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