Jujube witches’ broom phytoplasma effector SJP26 serves as a virulence factor to induce leaf yellowing

Leaf yellowing is among the common and easily identifiable symptoms that manifest during the invasion of phytoplasmas ( Candidatus Phytoplasma), leading to the repression of photosynthetic efficiency and losses in crop yield. Phytoplasma-secreted effectors attack several transcription factors to induce witches’ broom, phyllody and small leaves in jujube ( Ziziphus jujuba ). However, phytoplasma effector-induced leaf yellowing is not well understood. Here, the jujube witches’ broom (JWB) phytoplasma effector SJP26, which serves as a virulence factor to induce leaf yellowing in Arabidopsis thaliana and Nicotiana benthamiana , was characterized. The cytoplasmic- and nucleus-localized SJP26 encoded 136 amino acids (aa), containing one N-terminal signal peptide (1–33 aa), three α-helices and five β-sheets. SJP26 is highly expressed in JWB phytoplasma-infected leaves but is expressed at lower levels in the phloem and axillary bud. Heterologous expression of SJP26 without the signal peptide in A. thaliana resulted in a stronger ability to induce leaf yellowing than its homologues, including the SAP05 AYWB effector from the aster yellows phytoplasma strain witches’ broom, the SAP05 PnWBa effector from peanut witches’ broom phytoplasmas and the SAP05 WBDLa effector from witches’ broom disease of lime phytoplasmas. Transcriptomic analysis and pigment content measurements revealed that genes related to the chlorophyll biosynthetic pathway and chlorophyll content were significantly downregulated in the yellowed leaves of SJP26 transgenic N. benthamiana . Transformation of SJP26 in sour jujube calli confirmed the repression of gene expression related to chlorophyll biosynthesis and chlorophyll content, which were consistent with those in JWB phytoplasma-infected yellowed leaves. These findings provide insight into the mechanism underlying the effect of SJP26 on leaf yellowing in JWB phytoplasma-infected perennial woody fruit trees.