P53 mutation is an important cause of chemoresistance in colorectal cancer (CRC). The investigation and identification of the downstream targets and underlying molecular mechanism of chemoresistance induced by P53 abnormalities are therefore of great clinical significance. In this study, we demonstrated and reported for the first time that leucine-rich pentatricopeptide repeat-containing protein (LRPPRC) is a key functional downstream factor and therapeutic target for P53 mutation-induced chemoresistance. Due to its RNA binding function, LRPPRC specifically bound to the mRNA of multidrug resistance 1 (MDR1), increasing MDR1 mRNA stability and protein expression. In normal cells, P53 induced by chemotherapy inhibited the expression of LRPPRC via miR-34a and in turn reduced the expression of MDR1. However, chemotherapy-induced P53/miR-34a/LRPPRC/MDR1 signalling pathway activation was lost when P53 was mutated. Additionally, the accumulated LRPPRC and MDR1 promoted drug resistance. Most importantly, gossypol-acetic acid (GAA) was recently reported by our team as the first specific inhibitor of LRPPRC. In CRC cells with P53 mutation, GAA effectively induced degradation of the LRPPRC protein and reduced chemoresistance. Both in vivo and in vitro experiments revealed that combination chemotherapy with GAA and 5-fluorouracil (5FU) yielded improved treatment outcomes. In this study, we reported a novel mechanism and target related to P53-induced drug resistance and provided corresponding interventional strategies for the precision treatment of CRC.