Jujube Witches' Broom Phytoplasma Effectors SJP 1/2 Manipulate the ZjTCP 2‐ ZjTCP 1 Cascade to Repress Leaf Cell Proliferation in Jujube

Leaf development into the final size is spatiotemporally regulated by cell proliferation and expansion. The jujube witches' broom (JWB) phytoplasma effectors SJP1/2 induce small leaves with decreased cell numbers in jujube plants by promoting the accumulation of the CIN-TCP transcription factor ZjTCP2. However, the molecular mechanism underlying SJP1/2-mediated inhibition of leaf cell proliferation remains largely unknown. Here, we report that SJP1/2 repressed cell proliferation by manipulating ZjTCP2 to control the CYC/TB1-TCP transcription factor ZjTCP1-mediated pathway. Compared with the mature leaves of the control lines, overexpression of ZjTCP1 in Arabidopsis resulted in large leaves with increased cell numbers, which was in contrast to the findings in ZjTCP2-overexpressing transgenic jujube plants. Yeast one-hybrid, luciferase and reverse transcription-quantitative PCR assays revealed that ZjTCP2 directly targeted the ZjTCP1 promoter and downregulated its expression. Furthermore, SJP1 induced the degradation of ZjTCP1 protein. ZjTCP1 was highly expressed during early leaf growth and was positively correlated with cell proliferation-related genes, including ANGUSTIFOLIA3 (ZjAN3), AINTEGUMENTA (ZjANT) and B-type CYCLINS1;1 (ZjCYCB1;1). Further investigations confirmed that ZjTCP1 directly bound to and upregulated the expression of ZjAN3, ZjANT and ZjCYCB1;1, whereas these genes were significantly downregulated in both SJP1/2 and ZjTCP2 transgenic jujube plants and JWB phytoplasma-infected jujube trees compared with those in the controls. These results provide a new insight into the molecular mechanism of SJP1/2-induced small-leaf formation through the manipulation of the ZjTCP2-ZjTCP1 cascade in woody plants.