CpMADS4 and CpERF9 jointly regulate carotenoid synthesis related genes during papaya fruit ripening

Papaya carotenoid synthesis is a highly complex and coordinated biological metabolic process which involves the expression and regulation of hundreds to thousands of genes. However, little is known in the matter of the molecular modulation mechanism of papaya carotenoid synthesis in fruit ripening. In this investigation, we found that protein structure of MADS-box transcription factor CpMADS4 was highly similar to SlRIN, SlCMB1, AcRIN, PpMADS16 and CsMADS6, which played important roles in fruit development and carotenoid biosynthesis. CpMADS4 could not only directly interact with the promoters of CpPDS4, CpLCY-e and CpCHY-b, but also activate the expression of CpPDS4 and CpLCY-e. Protein-protein interaction experiment confirmed that CpMADS4 could interact with CpERF9, a suppressor in papaya fruit ripening. Moreover, CpERF9 repressed CpMADS4 in its transcriptional activation on CpLCY-e and CpPDS4. Overexpression of CpMADS4 promoted carotenoid synthesis and the expression of NtPDS and NtLCY-e, but overexpression of CpERF9 decreased the function of CpMADS4 on carotenoid synthesis in tobacco. Our results suggested that CpERF9 acted as a repressor of CpMADS4 in its transcriptional activation on carotenoid synthesis related genes promoters by interacting with CpMADS4. Taken together, our findings provided a new insight about MADS-box and ERF into the transcriptional regulation mechanism of carotenoid metabolism in papaya fruit ripening.