RESUMEN
Cellular autophagy is a widely-occurring conserved process for turning over damaged organelles or recycling cytoplasmic contents in cells. Although autophagy-related genes (ATGs) have been broadly identified from many plants, little is known about the potential function of autophagy in mediating plant growth and development, particularly in recycling cytoplasmic contents during seed development and germination. Castor bean (Ricinus communis) is one of the most important inedible oilseed crops. Its mature seed has a persistent and large endosperm with a hard and lignified seed coat, and is considered a model system for studying seed biology. Here, a total of 34 RcATG genes were identified in the castor bean genome and their sequence structures were characterized. The expressional profiles of these RcATGs were examined using RNA-seq and real-time PCR in a variety of tissues. In particular, we found that most RcATGs were significantly up-regulated in the later stage of seed coat development, tightly associated with the lignification of cell wall tissues. During seed germination, the expression patterns of most RcATGs were associated with the decomposition of storage oils. Furthermore, we observed by electron microscopy that the lipid droplets were directly swallowed by the vacuoles, suggesting that autophagy directly participates in mediating the decomposition of lipid droplets via the microlipophagy pathway in germinating castor bean seeds. This study provides novel insights into understanding the potential function of autophagy in mediating seed development and germination.
Asunto(s)
Proteínas Relacionadas con la Autofagia/genética , Perfilación de la Expresión Génica/métodos , Regulación de la Expresión Génica de las Plantas , Genoma de Planta/genética , Genómica/métodos , Ricinus communis/genética , Autofagia/genética , Proteínas Relacionadas con la Autofagia/clasificación , Proteínas Relacionadas con la Autofagia/metabolismo , Ricinus communis/metabolismo , Aceite de Ricino/metabolismo , Endospermo/genética , Endospermo/metabolismo , Germinación/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Gotas Lipídicas/metabolismo , Filogenia , Semillas/genética , Semillas/metabolismoRESUMEN
Protein-protein interaction (PPI) network maintains proper function of all organisms. Simple high-throughput technologies are desperately needed to delineate the landscape of PPI networks. While recent state-of-the-art yeast two-hybrid (Y2H) systems improved screening efficiency, either individual colony isolation, library preparation arrays, gene barcoding or massive sequencing are still required. Here, we developed a recombination-based 'library vs library' Y2H system (RLL-Y2H), by which multi-library screening can be accomplished in a single pool without any individual treatment. This system is based on the phiC31 integrase-mediated integration between bait and prey plasmids. The integrated fragments were digested by MmeI and subjected to deep sequencing to decode the interaction matrix. We applied this system to decipher the trans-kingdom interactome between Mycobacterium tuberculosis and host cells and further identified Rv2427c interfering with the phagosome-lysosome fusion. This concept can also be applied to other systems to screen protein-RNA and protein-DNA interactions and delineate signaling landscape in cells.