A single-base mutation in promoter of <i>CsTPR</i> enhances the negative regulation on mechanical-related leaf drooping in tea plants

Mechanical harvesting in the tea industry has become increasingly essential due to its advantages in increasing productivity and reducing labor costs. Leaf droopiness caused a high rate of broken leaves, hindering the mechanized harvesting quality. However, the underlying mechanisms remain unclear. We herein identified a quantitative trait locus, designated as <i>q10.3</i>, along with three lead single nucleotide polymorphisms (SNPs) located near a <i>TPR</i> gene (<i>TETRATRICOPEPTIDE REPEAT</i>), named <i>CsTPR</i>, through performing a genome-wide association study (GWAS) on 130 tea accessions. Integrated analysis of RNA-seq and ATAC-seq confirmed <i>CsTPR</i> as a droopiness-associated candidate gene at the transcriptional level. <i>CsTPR</i> was then proved to negatively regulate brassinosteroid-induced droopiness by using the <i>CsTPR</i>-silencing tea plant. Whole-genome sequencing (WGS) combined with genome walking further indicated that a single-base mutation (T-A) in the promoter of <i>CsTPR</i>. ChIP-seq revealed that this mutation occurred within the binding site, E-box, of CsBES1.2 on the <i>CsTPR</i> promoter. Notably, CsBES1.2 bound the E-box of <i>CsTPR</i> promoter to repress the expression of <i>CsTPR</i>, as demonstrated by chromatin immunoprecipitation quantitative polymerase chain reaction (ChIP-qPCR), electrophoretic mobility shift assays (EMSA), and transient assays. The single-base mutation strengthened the inhibitory effect of CsBES1.2 on the expression of <i>CsTPR</i> via enhancing the binding affinity to the E-box. Altogether, our findings suggest that CsTPR negatively regulates droopiness in tea plants under the transcriptional repression of CsBES1.2 and that a single-base mutation within E-box amplifies the suppression of CsBES1.2 on the expression of <i>CsTPR</i>.