Plant embryos are generated and develop in a stable and well-protected microenvironment surrounded by maternal tissue, which is vital for embryogenesis. However, the signaling mechanisms responsible for maternal tissue-to-proembryo communication are not well understood. Here, we report a pathway for maternal tissue-to-proembryo communication. We identify a DELLA protein, NtCRF1 ( NtCYS regulative factor 1), which regulates suspensor programmed cell death (PCD). NtCRF1 can bind to the promoter of NtCYS and regulate the suspensor PCD-switch module NtCYS-NtCP14 in response to gibberellin (GA). We confirm that GA 4 , as a primary signal triggering suspensor PCD, is generated in the micropylar endothelium by the transient activation of NtGA3oxs in the maternal tissue. Thus, we propose that GA is a maternal-to-proembryo communication signal that is decoded in the proembryo by a GID1-CRF1-CYS-CP14 signaling cascade. Using this mode of communication, maternal tissue precisely controls the embryonic suspensor PCD and is able to nurse the proembryo in a stage-dependent manner.
Embryonic suspensor in angiosperms is a short-lived structure that connects the embryo to surrounding maternal tissues, which is necessary for early embryogenesis. Timely degeneration via programed cell death is the most distinct feature of the suspensor during embryogenesis. Therefore, the molecular mechanism regulating suspensor cell death is worth in-depth study for embryonic development. However, this process can hardly be detected using conventional methods since early embryos are deeply embedded in the seed coats and inaccessible through traditional tissue section. Hence, it is necessary to develop a reliable protocol for terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling (TUNEL) analysis using limited living early embryos. Here, we provide a detailed protocol for the whole-mount detection of suspensor cell death using a TUNEL system in tobacco. This method is especially useful for the direct and rapid detection of the spatial-temporal characters of programed cell death during embryogenesis, as well as for the diminishment of the artifacts during material treatment by traditional methods.
Embryonic development is a key developmental event in plant sexual reproduction; however, regulatory networks of plant early embryonic development, particularly the effects and functional mechanisms of phospholipid molecules are still unknown due to the limitation of sample collection and analysis. We innovatively applied the microspore-derived in vitro embryogenesis of Brassica napus and revealed the dynamics of phospholipid molecules, especially phosphatidic acid (PA, an important second messenger that plays an important role in plant growth, development, and stress responses), at different embryonic developmental stages by using a lipidomics approach. Further analysis of Arabidopsis mutants deficiency of CDS1 and CDS2 (cytidinediphosphate diacylglycerol synthase, key protein in PA metabolism) revealed the delayed embryonic development from the proembryo stage, indicating the crucial effect of CDS and PA metabolism in early embryonic development. Decreased auxin level and disturbed polar localization of auxin efflux carrier PIN1 implicate that CDS-mediated PA metabolism may regulate early embryogenesis through modulating auxin transport and distribution. These results demonstrate the dynamics and importance of phospholipid molecules during embryo development, and provide informative clues to elucidate the regulatory network of embryogenesis.
Chloroplast is a semi-autonomous organelle with a double membrane structure, and its structural stability is the prerequisite for its correct function. Chloroplast development is regulated by known nuclear-encoded chloroplast proteins or proteins encoded within the chloroplast itself. However, the mechanism of chloroplast development is involved in other organelles remains largely unknown. Here, we report that a nuclear-located DEAD-box RNA helicase 13 (RH13) is essential for chloroplast development in Arabidopsis thaliana. RH13 is widely expressed in tissues and localized to the nucleolus. Homozygous rh13 mutant shows abnormal chloroplast structure and leaf morphogenesis. Proteomic analysis shows that the expression levels of photosynthesis related proteins in chloroplasts are reduced due to loss of RH13. Furthermore, RNA-sequencing and proteomics data reveals that the expression levels of these chloroplast-related genes are decreased which undergo alternative splicing events in rh13 mutant. Taken together, we propose that nucleolus-located RH13 is critical for chloroplast development in Arabidopsis.
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