ClPLD5, a phospholipase gene is involved in protection of cytoskeleton stability in pollen of self-imcompatible ‘Xiangshui’ lemon (Citrus limon)

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Self-incompatibility (SI) is an important mechanism for the prevention of self-fertilization in plants and is one of the major causes of seedlessness in citrus plants. Phospholipase D (PLD) is closely related to the growth and development of pollen, but the role of PLD in citrus plants during SI is poorly understood. In this study, we identified PLDs associated with SI in citrus plants, analyzed 15 C2-PLDs encoding homologs of the C2 domain in the genomes of sweet orange (Citrus sinensis) and clementine mandarin (Citrus clementina). We performed a phylogenic analysis to classify the PLD subfamilies of homologous genes from other species and found that the C2-PLDs of ‘Xiangshui’ lemon shared high homology with C2-PLDs from pear and Arabidopsis. Expression analysis revealed that ClPLD5 is probably involved in the regulation of pollen of ‘Xiangshui’ lemon during SI. ClPLD5 is enriched in pollen and functions as a pollen protection factor, as overexpression of ClPLD5 in self-incompatible pollen prevented the depolymerization of the actin cytoskeleton during SI. This work provides insights into the regulation of the cytoskeleton in pollen under SI challenge in citrus plants, in which PLD plays a protective role in incompatible pollen. A C2-PLD gene (ClPLD5) related to self-incompatibility was identified within the “Xiangshui” lemon C2-PLD family, and overexpression of the ClPLD5 gene in pollen increased the stability of the pollen cytoskeleton under self-incompatibility challenge.