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To better understand the molecular mechanism underlying of diapause in Antheraea pernyi (A. pernyi), we cloned a novel diapause-associated protein 3 (DAP3) gene from A. pernyi by reverse transcription-polymerase chain reaction (RT-PCR) and studied the biological functions. Sequence analysis revealed that this gene encodes 171 amino acids and has a conserved domain of Copper/Zinc superoxide dismutase (Cu/Zn-SOD). Western blot and qRT-PCR results showed that DAP3 was mainly expressed in the pupal stage, and gradually decreased as diapause development. DAP3 was also expressed in 1st and 5th instar larvae of A. pernyi. In tissues of 5th instar larvae of A. pernyi, DAP3 was mainly expressed in the epidermis, followed by the head, hemolymph and fat body. To identify the SOD activity of DAP3, we constructed a prokaryotic expression vector by inserting the coding region sequence into plasmid pET-28a (+) and obtained the purified rHIS-DAP3 fusion protein by Ni-NTA affinitive column. Importantly, we found the SOD activity of DAP3 fusion protein was approximately 0.6674 U/µg. To further confirm the SOD activity of DAP3 in vivo, we induced the oxidative stress model of pupae by UV irradiation. The results showed that both the mRNA and protein level of DAP3 significantly increased by UV irradiation. Furthermore, the SOD activity of the total lysate of pupae increased significantly at 10 min post UV irradiation and transiently returned to normal level afterwards. These results suggested that DAP3 might be a novel protein with SOD activity getting involved in regulation of diapause in A. pernyi.

Northern leaf blight, caused by the fungus Exserohilum turcicum (Pass.), is a major disease of maize (Zea mays L.). MicroRNAs (miRNAs) have been recently reported as gene expression regulators related to several stress responses; however, evidence of the role of miRNAs in plant response to biotic stresses is limited. In this study, the miRNA expression patterns in maize in response to E. turcicum stress were investigated using a plant miRNA microarray platform. A total of 118 miRNAs were detected in mock- and E. turcicum-inoculated leaves. Among these miRNAs, miR530, miR811, miR829, and miR845 were identified as new miRNAs in maize through a homology-based approach. The secondary structures and putative targets of these miRNAs were also predicted. In addition, four miRNAs were differentially regulated in response to E. turcicum: miR811, miR829, miR845, and miR408. The functional annotation of the predicted targets indicated that these stress-responsive miRNAs regulate metabolic, morphologic, and physiologic adaptations in maize seedlings at the post-transcriptional level. Four targets were negatively correlated with their corresponding miRNAs (miR811, miR829, and miR408). Furthermore, we have demonstrated for the first time that miR811 and miR829 confers a high degree of resistance to E. turcicum, which can be used in maize breeding programs.