Autophagy and vacuolar biogenesis during the nectary development.

MAIN CONCLUSION: Different autophagy pathways are a driver of vacuolar biogenesis and are development stage specific during the extrafloral nectary development in Citharexylum myrianthum. Plant autophagy plays an important role in various developmental processes such as seed germination, pollen maturation and leaf senescence. However, studies that address the evidence of autophagy and its role in the development of plant glands are scarce and largely restricted to laticifers. Regarding nectary, studies have repeatedly pointed to signs of degradation associated with the end of the secretory cycle, without exploring autophagy. Likewise, the relationship between autophagy and biogenesis of vacuoles remains an unexplored issue. In this study, using conventional and microwave fixation in association with ultracytochemical methods for transmission electron microscopy, we investigated the occurrence of autophagy and its implication in the differentiation of extrafloral nectary in Citharexylum myrianthum (Verbenaceae) under natural conditions, focusing on the vacuole biogenesis. We described a variety of vacuole types associated with the stage of nectary epidermis development, which differs with respect to origin, function and nature of the products to be stored. Three distinct autophagy pathways were detected: macroautophagy, microautophagy (both restricted to the undifferentiated epidermal cells, at the presecretory stage) and megaautophagy (circumscribed to the differentiated epidermal cells, at the postsecretory stage). Our study clearly demonstrated that the vacuole variety and autophagy processes in the nectary epidermal cells are development specific. This study highlights the role of autophagy in vacuole biogenesis and its implications for the development of nectary and opens new venues for future studies on regulation mechanisms for autophagy in plant secretory structures under normal conditions.

Structural associations between organelle membranes in nectary parenchyma cells.

MAIN CONCLUSION: The close association between membranes and organelles, and the intense chloroplast remodeling in parenchyma cells of extrafloral nectaries occurred only at the secretion time and suggest a relationship with the nectar secretion. Associations between membranes and organelles have been well documented in different tissues and cells of plants, but poorly explored in secretory cells. Here, we described the close physical juxtaposition between membranes and organelles, mainly with chloroplasts, in parenchyma cells of Citharexylum myrianthum (Verbenaeceae) extrafloral nectaries under transmission electron microscopy, using conventional and microwave fixation. At the time of nectar secretion, nectary parenchyma cells exhibit a multitude of different organelle and membrane associations as mitochondria-mitochondria, mitochondria-endoplasmic reticulum, mitochondria-chloroplast, chloroplast-nuclear envelope, mitochondria-nuclear envelope, chloroplast-plasmalemma, chloroplast-chloroplast, chloroplast-tonoplast, chloroplast-peroxisome, and mitochondria-peroxisome. These associations were visualized as amorphous electron-dense material, a network of dense fibrillar material and/or dense bridges. Chloroplasts exhibited protrusions variable in shape and extension, which bring them closer to each other and to plasmalemma, tonoplast, and nuclear envelope. Parenchyma cells in the pre- and post-secretory stages did not exhibit any association or juxtaposition of membranes and organelles, and chloroplast protrusions were absent. Chloroplasts had peripheral reticulum that was more developed in the secretory stage. We propose that such subcellular phenomena during the time of nectar secretion optimize the movement of signaling molecules and the exchange of metabolites. Our results open new avenues on the potential mechanisms of organelle contact in parenchyma nectary cells, and reveal new attributes of the secretory cells on the subcellular level.