Sunflower pollen is a natural nutritious food with a long history and multiple functions, however, the main chemical components apart from flavonoids and their biosynthesis processes have not been thoroughly investigated. In this study, seven hydroxycinnamic acid amides (HCAAs) (1-7) abundant in sunflower pollen were isolated and identified as one type of the pollen's main chemicals. For a comprehensive understanding of HCAA biosynthesis in Helianthus annuus flowers, RNA-seq, metabolomics, and key genes related to biosynthesis in the sunflower were studied. A large number of compounds at different sunflower growth stages (the 7th, 14th, 21st, and 28th days) and high expression levels of related genes in the transcriptome were detected. A molecular network was constructed to clarify the synthetic pathway of HCAAs, which revealed high transcriptional levels of spermidine hydroxycinnamoyl transferase genes (HaSHT2795 and HaSHT2436) in 14-21-days-old flowers. HaSHT2795 enzymes catalyze tri-coumaroylspermidine formation, and virus-induced gene silencing to inhibit HaSHT2795 and HaSHT2436 could significantly reduce the synthesis of hydroxycinnamic acid amides in sunflower pollen. HCAAs were inferred to be related to the formation of pollen walls and the health effects of pollen. Analyzing HCAA biosynthesis and accumulation in H. annuus pollen will be helpful to understand the functions of HCAAs in the development of pollen and its nutritional value.
Keywords: Biosynthesis; Co-expression network; Metabolomics; Spermidine hydroxycinnamoyl transferase; Sunflower pollen; ·Hydroxycinnamic acid amides.
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