Gibberellin regulates the synthesis of stone cells in 'Nanguo' pear via the PuMYB91-PuERF023 module

Stone cells are one of the limiting factors affecting pear fruit quality and commodity value. The formation of stone cell is highly correlated with lignin deposition. However, the molecular mechanism of stone cell formation and regulation is still unclear. Here, we observed that exogenous application of GA significantly inhibited the formation of stone cells and also decreased the content of lignin in 'Nanguo' (Pyrus ussuriensis) pear fruits. The key gene PuPRX73 involved in the lignin synthesis pathway was further identified using RT-PCR, and GA-treatment significantly inhibited the expression of PuPRX73. Overexpression or silencing of PuPRX73 in pear fruits significantly increases or decreases the content of stone cells and lignin. We identified the transcription factors PuMYB91 and PuERF023 using mRNA-seq and their expression was significantly decreased after GA-treatment. Transient overexpression of PuMYB91 and PuERF023 promotes lignin and stone cells content in pear fruits, while silencing of PuMYB91 and PuERF023 led to the opposite results and inhibited the expression of PuPRX73. Yeast one-hybrid (Y1H) and GUS activity analysis revealed that PuMYB91 and PuERF023 directly bind and activate the PuPRX73 promoter, and co-transfection of PuMYB91 and PuERF023 in Nicotiana benthamiana leaves further promoted the promoter activity of PuPRX73. Furthermore, we found that PuMYB91 interacted with PuERF023 in vitro by using Yeast two-hybrid assays (Y2H). In conclusion, our results revealed that exogenous GA-treatment inhibits stone cell production by suppressing the expression of PuMYB91 and PuERF023 in pear fruits.