Comprehensive studies of the serine carboxypeptidase-like (SCPL) gene family in Carya cathayensis revealed the roles of SCPL4 in epigallocatechin-3-gallate (EGCG) synthesis and drought tolerance

Hickory (Carya cathayensis) nuts are rich in epigallocatechin-3-gallate (EGCG) with multiple health functions. EGCG also regulates plant growth, development and stress responses. However, research on the synthesis mechanism of EGCG and its function in hickory is currently limited. Herein, 44 serine carboxypeptidase-like (SCPL) members were identified from the hickory genome and classified into three major categories: SCPL-I, SCPL-II, and SCPL-III. In the CcSCPLs-IA branch, CcSCPL3/4/5/8/9/11/13 showed differential expression patterns in various tissues, especially with relatively high expression levels in plant roots, female flowers and seed coat. These proteins have a catalytic triad composed of serine (Ser), aspartic acid (Asp) and histidine (His). Ser-His in the triad and arginine (Arg) mediated the docking of CcSCPL3/4/5/11 with 1-O-galloyl-β-d-glucose (βG) and epigallocatechin (EGC), whereas the Asp of the triad did not. CcSCPL4 was further confirmed to promote the synthesis of EGCG in tobacco leaves. CcSCPL4 may function as monomer and be mainly localized within cellular structures outside the nucleus. Notably, the expression level of CcSCPL4 significantly changed after drought, cold, and salt stress, with the highest expression level under drought stress. Meanwhile CcSCPL4 over-expression could enhance the drought resistance of Saccharomyces cerevisiae and Arabidopsis. This study elucidates key enzymes for EGCG synthesis and their role in drought resistance, providing insights into the EGCG synthesis pathway and molecular breeding of hickory in future.