The morphology and physiology of diaspores play crucial roles in determining the fate of seeds in unpredictable habitats. In some genera of the Brassicaceae different types of diaspores can be found. Lepidium appelianum produces non-dormant seeds within indehiscent fruits while in L. campestre dormant seeds are released from dehiscent fruits. We investigated whether the allocation of relevant defence compounds into different tissues in different Lepidium species may be related to the diverse dispersal strategy (indehiscent and dehiscent) and seed physiology (non-dormant and dormant). Total glucosinolate concentration and composition were analysed in immature and mature seeds and pericarps of L. appelianum and L. campestre using high-performance liquid chromatography. Moreover, for comparison, transgenic RNAi L. campestre lines were used that produce indehiscent fruits due to silencing of LcINDEHISCENCE, the INDEHISCENCE ortholog of L. campestre. Total glucosinolate concentrations were lower in immature compared to mature seeds in all studied Lepidium species and transgenic lines. In contrast, indehiscent fruits of L. appelianum maintained their total glucosinolate concentration in mature pericarps compared to immature ones, while in dehiscent L. campestre and in indehiscent RNAi-LcIND L. campestre a significant decrease in total glucosinolate concentrations from immature to mature pericarps could be detected. Indole glucosinolates were detected in lower abundance than the other glucosinolate classes (aliphatic and aromatic). Relatively high concentrations of 4methoxyindol-3-ylmethyl glucosinolate were found in mature seeds of L. appelianum compared to other tissues, while no indole glucosinolates were detected in mature diaspores of L. campestre. The diaspores of the latter species may rather depend on aliphatic and aromatic glucosinolates for long-term protection. The allocation patterns of glucosinolates correlate with the morpho-physiologically distinct fruits of L. appelianum and L. campestre and
20The morphology and physiology of diaspores play crucial roles in determining the fate of 21 seeds in unpredictable habitats. In some genera of the Brassicaceae different types of 22 diaspores can be found. Lepidium appelianum produces non-dormant seeds within 23 indehiscent fruits while in L. campestre dormant seeds are released from dehiscent fruits. 24These different diaspore types offer an excellent model system to analyse the allocation 25 of relevant defence compounds into different tissues, which may maximise diaspore 26 fitness. Total glucosinolate concentration and composition were analysed in immature and 27 mature seeds and pericarps of L. appelianum and L. campestre using high-performance 28 liquid chromatography. Moreover, transgenic RNAi L. campestre lines were used for 29 comparison that produce indehiscent fruits due to silencing of LcINDEHISCENCE, the 30 INDEHISCENCE ortholog of L. campestre. Total glucosinolate concentrations were lower 31 in green compared to mature seeds in all studied Lepidium species and transgenic lines. 32In contrast, indehiscent fruits of L. appelianum maintained their total glucosinolate 33 concentration in mature pericarps compared to green ones, while in dehiscent L. 34 campestre and in indehiscent RNAi-LcIND L. campestre a significant decrease in total 35 glucosinolate concentrations from green to mature pericarps could be detected. 36Regarding the distribution of glucosinolate classes, high concentrations of 4-37 methoxyindol-3-ylmethyl glucosinolate were found in mature seeds of L. appelianum, 38 while no indole glucosinolates were detected in mature diaspores of L. campestre. The 39 diaspores of the latter species may rather depend on aliphatic and aromatic glucosinolates 40 for long-term protection. The allocation patterns of glucosinolates correlate with the 41 morpho-physiologically distinct fruits of L. appelianum and L. campestre and may be 42 explained by the distinct dispersal strategies and the dormancy status of both species. 43 in the diaspores using transgenic RNAi-LcIND L. campestre. Finally, we tested the 107 longevity in the seedbank for the wild type plants of both species. We discuss the 108 allocation of GSLs in the diaspores of L. appelianum and L. campestre in relation to their 109 dispersal strategy and their natural seedbank persistence and dormancy cycle. 110 Materials and Methods111 Seed sources 112 Seeds of Lepidium appelianum (KM 1754; obtained from J Gaskin, USDA, Fremont 113 County, Wyoming, USA) and wild type L. campestre (KM 96; obtained from Botanical 114 Garden, University of Zürich) were collected from mass propagations in the Botanical 115 Garden, Osnabrueck University, Germany, in 2014 to 2015. Seeds of the transgenic 116 Lepidium campestre, in which RNAi silencing of LcIND (pFGC5941:LcINDa) resulting in 117 indehiscent fruits is established (see details of transformation and plant cultivation 118 procedures in [24]), were collected from plants cultivated at
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