Identification of autophagy-related genes ATG4 and ATG8 from wheat (Triticum aestivum L.) and profiling of their expression patterns responding to biotic and abiotic stresses.

The genes coding for wheat ATG4 and ATG8 were cloned and their roles in autophagy were verified. Implications of ATG4/ATG8 in wheat responses to stresses were suggested by expression profiling. Autophagy-related proteins ATG4 and ATG8 are crucial for autophagy biogenesis. ATG4 processes ATG8 precursor to expose its C-terminal glycine for phosphatidyl ethanolamine (PE) lipidation. ATG8, in the form of ATG8-PE adduct, functions in the organization dynamics of autophagic membranes. Here, we report the identification of two/nine members of the ATG4/ATG8 family from common wheat (Triticum aestivum L.). Expression of each wheat ATG4/ATG8 could complement the autophagy activity of yeast atg4/atg8 mutant cells. GFP fusion proteins of ATG8s, especially of ATG8s with innate C-terminal-exposed glycines, localized to punctate autophagic membranes. Both of purified ATG4s could cleave ATG8s in vitro, but they had different activities and different preferences for ATG8 substrates. Two times of transcript accumulation, an early one and a late one, of ATG4s/ATG8s were detected in the early phases of the Pm21- and Pm3f-triggered wheat incompatible reactions to the powdery mildew causal fungus Blumeria graminis f. sp. tritici (Bgt), and fluorescence microscopy also revealed a Bgt-induced enhanced wheat autophagy level in the Pm21-triggered incompatible reaction. Only one time of Bgt-induced transcript accumulation of ATG4s/ATG8s, corresponding to but much higher than the late one in incompatible reactions, was detected in a susceptible line isogenic to the Pm21 resistance line. These results suggested positive roles of ATG4/ATG8-associated autophagy process in the early stage and possible negative roles in the late stage of wheat immunity response to Bgt. In addition, expression of wheat ATG4s/ATG8s was also found to be upregulated by abiotic stress factors and distinctively regulated by different phytohormones.