PbbHLH41 negatively regulates iron deficiency tolerance in pear

Iron (Fe) is a trace element that is essential for plant growth and development. Thus, Fe-deficiency stress seriously affects plant growth, causing symptoms such as chlorosis and growth retardation. Furthermore, Fe-deficiency stress substantially reduces the yield and quality of pear fruits. In this study, we found that the bHLH transcription factor PbbHLH41 was considerably downregulated under Fe-deficiency conditions. The PbbHLH41-overexpressed tomato plants were obtained, and functional assays showed that PbbHLH41 overexpression substantially reduced the tomato plants' Fe-deficiency tolerance. Moreover, we generated PbbHLH41-knockout calli using gene-editing technology, and compared to wild-type calli, the PbbHLH41-knockout calli exhibited notably enhanced tolerance to Fe-deficiency stress. Further studies found that PbbHLH41 substantially downregulated the expression of Fe uptake-related genes, including PbAHA12, PbFRO4, PbFRO2-like, and PbIRT1. Dual-luciferase assays and yeast one-hybrid assays demonstrated that PbbHLH41 can bind to the promoters of PbFRO2-like and PbIRT1 and inhibit their expression. In conclusion, this study demonstrates that PbbHLH41 negatively regulates Fe-deficiency stress tolerance, thereby providing genetic information and a theoretical basis for pear rootstock breeding to enhance their tolerance to Fe-deficiency stress.