Antepenultimate residue at the C-terminus of NADPH oxidase RBOHD is critical for its function in the production of reactive oxygen species in Arabidopsis / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Production of reactive oxygen species (ROS) is a conserved immune response primarily mediated by NADPH oxidases (NOXs), also known in plants as respiratory burst oxidase homologs (RBOHs). Most microbe-associated molecular patterns (MAMPs) trigger a very fast and transient ROS burst in plants. However, recently, we found that lipopolysaccharides (LPS), a typical bacterial MAMP, triggered a biphasic ROS burst. In this study, we isolated mutants defective in LPS-triggered biphasic ROS burst (delt) in Arabidopsis, and cloned the DELT1 gene that was shown to encode RBOHD. In the delt1-2 allele, the antepenultimate residue, glutamic acid (E919), at the C-terminus of RBOHD was mutated to lysine (K). E919 is a highly conserved residue in NADPH oxidases, and a mutation of the corresponding residue E568 in human NOX2 has been reported to be one of the causes of chronic granulomatous disease. Consistently, we found that residue E919 was indispensable for RBOHD function in the MAMP-induced ROS burst and stomatal closure. It has been suggested that the mutation of this residue in other NADPH oxidases impairs the protein's stability and complex assembly. However, we found that the E919K mutation did not affect RBOHD protein abundance or the ability of protein association, suggesting that the residue E919 in RBOHD might have a regulatory mechanism different from that of other NOXs. Taken together, our results confirm that the antepenultimate residue E is critical for NADPH oxidases and provide a new insight into the regulatory mechanisms of RBOHD.

The ethylene signal mediates induction of GmATG8i in soybean plants under starvation stress.

In higher plants, autophagy-related genes (ATGs) appear to play important roles in development, senescence, and starvation responses. Hormone signals underlying starvation-induced gene expression are involved in the expression of ATGs. An effect of starvation stress on the expression of ATGs and ethylene-related genes in young seedlings of soybean (Glycine max [L.] Merr. cv. Fukuyutaka) was analyzed. Reverse transcription-polymerase chain reaction (RT-PCR) showed that the expression levels of GmATG8i and GmATG4 increase in a starvation medium, but at a null or marginal level in a sucrose/nitrate-rich medium. The expression of GmACC synthase and GmERF are also upregulated in the starvation medium. In addition, immunoblot revealed that ethylene insensitive 3 (Ein3), an ethylene-induced transcription factor are accumulated in seedlings subjected to severe starvation stress. These results indicate that starvation stress stimulates the expression of GmATG8i and ethylene signal-related genes. Since the ethylene signal is involved in senescence and various environmental stresses, it is possible that starvation stress-induced autophagy is partly mediated by the ethylene signaling.