Identification of the 2AP Regulatory Gene CnProDH in Aromatic Coconut and Screening of Its Regulatory Factors

Aromatic coconut is a special variety of coconut. Its unique “pandan-like” aroma has won it great popularity among consumers, endowing it with considerable market potential. In our previous study, 2-acetyl-1-pyrroline (2AP), which serves as the main source of the “pandan-like” aroma in aromatic coconut, was found to exhibit significant variation among distinct aromatic coconut individuals. Now, the regulatory mechanism of 2AP has been clarified in fragrant rice, and the ProDH gene is the key gene for 2AP regulation. To further understand the regulation mechanism of 2AP content in aromatic coconut, we cloned and identified the CnProDH gene, the key gene of 2AP regulation in aromatic coconut. The results showed that the CnProDH gene had the typical ProDH structural domain, and its full-length sequence is 23,667 bp, containing 5 exons and a coding sequence (CDS) of 1599 bp. The CnProDH gene encodes a protein that possesses a β8α8 barrel structure, consisting of 532 amino acids (aa), with a molecular mass of 58,076.63 kDa and an isoelectric point of 7.11. To further understand the regulatory mechanism of CnProDH in aromatic coconut, we also constructed a yeast one-hybrid (Y1H) library for aromatic coconut. Through the Y1H experiment, combined with the prediction and analysis of cis-acting elements in the promoter of the CnProDH gene, three possible regulatory factors, including CnYABBY2, CnSAP8, and CnBRD3, were identified. These findings provide a molecular basis for clarifying and solving the problem of variations in 2AP content across different aromatic coconuts.