Self-incompatible-like response induces alterations of actin cytoskeleton and cytosolic calcium in the Citrus pollen tube of 'Banpeiyu' and 'Hyuganatsu'

Self-incompatibility (SI) in Citrus strongly prevents self-pollination and promotes outcrossing. Even though it has been confirmed that the S-allele is a key gene in controlling the recognition between pollen and pistil in Rosaceae, Solanaceae and Papaveraceae, this has not been cloned in Citrus. A citrus mature pollen culture system was established in our lab in order to culture 'Banpeiyu' and 'Hyuganatsu' pollen in vitro. Stylar crude protein extracts were used to treat citrus pollen in order to induce the SI-like response in the pollen tube. We observed the alteration of the pollen tube shape, including tip twisting and content leakage. Programmed cell death (PCD), as a normal cellular process that eliminates unwanted cells, was reported in previous studies of incompatible pollen tubes, together with an increase in hydrogen peroxide (H2O2), depolymerization of actin skeleton, the collapse of mitochondria, and leakage of cytochrome c. In our study, we investigated the cytosolic calcium ion concentration ([Ca2+](cyt)) changes and identified alterations in the actin cytoskeleton in the pollen tube after exposure to the incompatible treatment. The [Ca2+](cyt) content increased after incompatible treatment, but there was no obvious difference between the compatible-treatment group and the control. Punctate foci of actin filaments were found in the incompatible-treated pollen tubes; the rates of punctate foci were significantly higher than in the compatible-treated pollen tubes in both 'Banpeiyu' and 'Hyuganatsu'. The results suggested that these alterations in the pollen tubes were directly induced by the SI-like response, and might cause pollen tube PCD.