CsTCP14-CsIAA4 module-mediated repression of auxin signaling regulates citrus somatic embryogenesis

Somatic embryogenesis (SE) is an important in vitro regeneration approach for plants, especially in biotechnological manipulations. However, SE capability is difficult to modulate and deteriorates over time. Glycerol medium is effective in SE induction of citrus, while the mechanisms remain unclear. We found that auxin signaling reduced soon after the citrus embryogenic callus (EC) was transferred to glycerol medium, and the expression of CsTCP14 and AUX/IAA gene CsIAA4 was induced by glycerol. Overexpression of CsIAAm that encodes a stable indole-3-acetic acid (IAA) protein suppressed auxin signaling in EC and enhanced SE. CsTCP14 bound to the promoter of CsIAA4 and activate CsIAA4 expression in EC with strong SE competence. Overexpression of CsTCP14 activated CsIAA4 expression and reduced auxin signaling in citrus EC, and thus enhanced SE. Application of exogenous IAA inhibits SE, while the auxin signaling inhibitor p-chlorophenoxyisobutyric acid (PCIB) enhances SE in citrus. The SE enhancement effect of CsIAA4 and CsTCP14 overexpression on EC was alleviated by exogenous IAA, but reinforced by PCIB. We uncover the regulatory pathway of CsTCP14-CsIAA4 module-mediated repression of auxin signaling in glycerol-induced citrus SE, which deepens our understanding of SE mechanisms in plants and supports modulation of SE in citrus breeding via biotechnological approaches.