Methods for analysis of autophagy in plants

The plant vacuole is a major site for the breakdown and recycling of cellular macromolecules. Cytoplasmic components destined for degradation are delivered to the vacuole in vesicles termed autophagosomes, and the breakdown products are transported back into the cytosol for reuse, with the overall process termed autophagy. In plants, autophagy is required for nutrient remobilization and recycling during senescence and nutrient deficiency, for clearance of protein aggregates and damaged organelles during environmental stress, for pathogen defense, and for general cellular maintenance under normal growth conditions. There is growing interest in autophagy in plants due to the wide range of processes in which it functions. While much of the work thus far has used the model plant Arabidopsis thaliana, autophagy is now under investigation in a number of other plants, particularly in economically important crop species. Here, I discuss methods for assessing autophagy activity in plant cells. Microscopic and biochemical assays are described, along with ways to distinguish the steady-state number of autophagosomes from flux through the autophagic pathway. Some deficiencies still exist in plant autophagy analysis, and there is a particular need for more accurate methods of quantifying autophagic flux in plants. (C) 2014 Elsevier Inc. All rights reserved.