NAC (no apical meristem (NAM), Arabidopsis thaliana transcription activation factor (ATAF1/2) and cup shaped cotyledon (CUC2)) transcription factors play crucial roles in plant development and stress responses. Nevertheless, to date, only a few reports regarding stress-related NAC genes are available in Malus baccata (L.) Borkh. In this study, the transcription factor MbNAC25 in M. baccata was isolated as a member of the plant-specific NAC family that regulates stress responses. Expression of MbNAC25 was induced by abiotic stresses such as drought, cold, high salinity and heat. The ORF of MbNAC25 is 1122 bp, encodes 373 amino acids and subcellular localization showed that MbNAC25 protein was localized in the nucleus. In addition, MbNAC25 was highly expressed in new leaves and stems using real-time PCR. To analyze the function of MbNAC25 in plants, we generated transgenic Arabidopsis plants that overexpressed MbNAC25. Under low-temperature stress (4 °C) and high-salt stress (200 mM NaCl), plants overexpressing MbNAC25 enhanced tolerance against cold and drought salinity conferring a higher survival rate than that of wild-type (WT). Correspondingly, the chlorophyll content, proline content, the activities of antioxidant enzymes superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) were significantly increased, while malondialdehyde (MDA) content was lower. These results indicated that the overexpression of MbNAC25 in Arabidopsis plants improved the tolerance to cold and salinity stress via enhanced scavenging capability of reactive oxygen species (ROS).

Plant-specific WRKY transcription factors were involved in stress responses and ABA signaling. In the present study, a WRKY gene is isolated from Malus baccata (L.) Borkh and designated as MbWRKY3. Subcellular localization revealed that MbWRKY3 was localized onto nucleus. The MbWRKY3 expression levels were up-regulated by salinity, drought, and ABA treatments in M. baccata. When MbWRKY3 was introduced into tobaccos, it improved drought stress tolerance in transgenic plants. Compared to WT, the transgenic tobaccos had the higher levels of relative water content, and proline and chlorophyll contents, decreased levels of electrolyte leakage, MDA, and H₂O₂, increased activities of the reactive oxygen species-related enzymes (SOD, CAT, and POD), and greater up-regulations of the corresponding genes (NtSOD, NtCAT, and NtPOD), especially when dealt with drought stress. These results suggest that MbWRKY3 gene plays a positive regulatory role in drought stress response.

Plant-specific WRKY transcription factors are widely involved in abiotic stress responses. In this study, a WRKY gene was isolated from Malus baccata (L.) Borkh and designated it as MbWRKY2. Subcellular localization showed that MbWRKY2 was localized in the nucleus. The expression levels of MbWRKY2 were up-regulated by abiotic stresses in M. baccata. When MbWRKY2 was introduced into tobacco, it improved drought stress tolerance in transgenic plants. The transgenic tobaccos had the higher contents of chlorophyl, proline, relative water content, AsA, and GSH, increased activities of CAT, APX, SOD, and POD, and decreased levels of MDA, H₂O₂, and electrolyte leakage than wild-type, especially when dealt with dehydration treatment. Moreover, the MbWRKY2-OE plants enhanced the expression of oxidative stress response and stress-related genes involved in osmotic adjustment and membrane protection. These results suggest that MbWRKY2 gene plays a positive regulatory role in drought stress response.

Apple, as one of the most important economic forest tree species, is widely grown in the world. Abiotic stress, such as low temperature and high salt, affect apple growth and development. Ethylene response factors (ERFs) are widely involved in the responses of plants to biotic and abiotic stresses. In this study, a new ethylene response factor gene was isolated from Malus baccata (L.) Borkh and designated as MbERF11. The MbERF11 gene encoded a protein of 160 amino acid residues with a theoretical isoelectric point of 9.27 and a predicated molecular mass of 17.97 kDa. Subcellular localization showed that MbERF11 was localized to the nucleus. The expression of MbERF11 was enriched in root and stem, and was highly affected by cold, salt, and ethylene treatments in M. baccata seedlings. When MbERF11 was introduced into Arabidopsis thaliana, it greatly increased the cold and salt tolerance in transgenic plant. Increased expression of MbERF11 in transgenic A. thaliana also resulted in higher activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), higher contents of proline and chlorophyll, while malondialdehyde (MDA) content was lower, especially in response to cold and salt stress. Therefore, these results suggest that MbERF11 probably plays an important role in the response to cold and salt stress in Arabidopsis by enhancing the scavenging capability for reactive oxygen species (ROS).