Functional characterization of CqCYP86A1a and CqCYP86A1b in root suberin biosynthesis in Chenopodium quinoa.

CYP86A1 encodes a fatty acid ω-hydroxylase that is required for suberin monomer biosynthesis and plays an essential role in root suberization in Arabidopsis, rice and tomato. However, whether CYP86A1 function is conserved in Chenopodium quinoa, a typical halophyte, remains unclear. In this study, we identified two CYP86A1 homologs from quinoa, CqCYP86A1a and CqCYP86A1b, both of which were primarily expressed in quinoa roots and highly induced by salt stress. Further analysis showed that overexpression of either CqCYP86A1a or CqCYP86A1b completely rescued the defective root suberization phenotype of Arabidopsis cyp86a1 seedlings, whereas CqCYP86A1b overexpression further impaired suberin biosynthesis in mature cyp86a1 roots. Consistently, transient expression in Nicotiana benthamiana revealed that CqCYP86A1a exhibited much stronger activity in promoting suberin biosynthesis than CqCYP86A1b. Sequence comparison showed that three variations in CqCYP86A1b, V290F, K341N and M504T, are located in conserved substrate recognition sites. Further site-directed mutagenesis combined with transient expression in tobacco demonstrated that V290F and M504T variations significantly affected CqCYP86A1b activity and protein abundance. Together, these findings expand our understanding of CYP86A1 function and provide new evidence that CYP86A1-mediated suberin biosynthesis is functionally conserved in halophytes.