Synergistic impact of MdERF2 and MdPUB17 on the biosynthesis of wax in apple epidermis

The synergistic regulatory effect of the ethylene transcription factor MdERF2 and ubiquitin ligase MdPUB17 on apple (Malus domestica) epidermal wax was examined by transferring the pRI101-MdPUB17-MdERF2 dual overexpression vector (PUB17-ERF2), the empty vector (pRI101), the pRI101-MdPUB17 overexpression vector (PUB17), and the pRI101-MdERF2 overexpression vector (ERF2) into Agrobacterium tumefaciens, respectively, to infect apple callus and fruits with water as the control (CK). The levels of expression of the genes related to the biosynthesis, transport, composition, content, and structure of wax in the callus and/or fruits were studied under different treatments. The synergistic treatment of PUB17-ERF2 resulted in a decrease in the expression levels of MdCER1, MdCER6, MdLACS2, MdWSD1, MdABCG11, MdPAS2, MdFATB, and MdKASII genes as induced by the sole treatment of ERF2. Moreover, in the treatment of PUB17-ERF2, the mass distribution density of the wax was observed to be intermediate between the ERF2 and PUB17 treatments. Furthermore, ERF2 was found to increase the contents of alkanes, alcohols, and ketones, while significantly decreasing the contents of fatty acids and esters. In contrast, PUB17 responded oppositely. When treated with PUB17-ERF2, the effects of PUB17 and ERF2 were observed to counteract each other, which resulted in intermediate levels of these compounds. Additionally, the fruit in the ERF2, PUB17 and PUB17-ERF2 treatments had a different waxy microstructure. Overall, the findings indicate that both ERF2 and PUB17 have an impact on the gene expression, wax composition, content, and microstructure in apple epidermis. Importantly, the co-expression of MdPUB17 and MdERF2 demonstrates their synergistic regulation of the biosynthesis of wax in the apple epidermis.