Actin Dynamics in Papilla Cells of Brassica rapa during Self- and Cross-Pollination

Abstract

The self-incompatibility system of the plant species Brassica is controlled by the S-locus, which contains S-RECEPTOR KINASE (SRK) and S-LOCUS PROTEIN11 (SP11). SP11 binding to SRK induces SRK autophosphorylation and initiates a signaling cascade leading to the rejection of self pollen. However, the mechanism controlling hydration and germination arrest during self-pollination is unclear. In this study, we examined the role of actin, a key cytoskeletal component regulating the transport system for hydration and germination in the papilla cell during pollination. Using rhodamine-phalloidin staining, we showed that cross-pollination induced actin polymerization, whereas self-pollination induced actin reorganization and likely depolymerization. By monitoring transiently expressed green fluorescent protein fused to the actin-binding domain of mouse talin, we observed the concentration of actin bundles at the cross-pollen attachment site and actin reorganization and likely depolymerization at the self-pollen attachment site; the results correspond to those obtained by rhodamine-phalloidin staining. We further showed that the coat of self pollen is sufficient to mediate this response. The actin-depolymerizing drug cytochalasin D significantly inhibited pollen hydration and germination during cross-pollination, further emphasizing a role for actin in these processes. Additionally, three-dimensional electron microscopic tomography revealed the close association of the actin cytoskeleton with an apical vacuole network. Self-pollination disrupted the vacuole network, whereas crosspollination led to vacuolar rearrangements toward the site of pollen attachment. Taken together, our data suggest that self-and cross-pollination differentially affect the dynamics of the actin cytoskeleton, leading to changes in vacuolar structure associated with hydration and germination.

S-RNase disrupts tip-localized reactive oxygen species and induces nuclear DNA degradation in incompatible pollen tubes of Pyrus pyrifolia

“…Endomembrane breakdown in incompatible pollen tubes was found in Nicotiana (Goldraij et al, 2006) and also in Brassica, where vacuolar disruption in incompatible stigmatic papilla cells and rejection is triggered by SRK (S-locus receptor kinase) (Iwano et al, 2007). Actin depolymerization was also detected in stigmatic papilla cells (Iwano et al, 2007). These data indicate a potential link among the different SI systems.…”

“…The mechanisms involved in inhibition of pollen by the pear SI response shared some features with the mechanistically different SI in the poppy, where S-protein (recently renamed PrsS) triggers a Ca 2+ -dependent signal cascade, including protein phosphorylation, depolymerization of actin cytoskeleton and microtubules, and PCD (Geitmann et al, 2000;Staiger and Franklin-Tong, 2003;Thomas and Franklin-Tong, 2004;McClure and Franklin-Tong, 2006;Thomas et al, 2006;Poulter et al, 2008). Endomembrane breakdown in incompatible pollen tubes was found in Nicotiana (Goldraij et al, 2006) and also in Brassica, where vacuolar disruption in incompatible stigmatic papilla cells and rejection is triggered by SRK (S-locus receptor kinase) (Iwano et al, 2007). Actin depolymerization was also detected in stigmatic papilla cells (Iwano et al, 2007).…”

A Pollen Coat–Inducible Autoinhibited Ca2+-ATPase Expressed in Stigmatic Papilla Cells Is Required for Compatible Pollination in the Brassicaceae

“…The pollen surface is covered with exine and a protein-and lipid-rich substance called the pollen coat. The pollen coat alone from cross-pollen grains induced actin polymerization, while self-pollen coat alone induced the disappearance of actin bundles (Iwano et al, 2007). Furthermore, electron microscopic observation in Brassica oleracea revealed that applying the isolated pollen coat to the stigma caused cell wall expansion (Elleman and Dickinson, 1996).…”

“…Therefore, we speculated that unknown molecules in the pollen grain elicit a Ca 2+ -dependent signaling pathway in the papilla cells and that preexisting molecules in the papilla cells keep intracellular [Ca 2+ ] at low levels during compatible pollination, while extracellular [Ca 2+ ] is high. Cell biological analysis of B. rapa showed that cross-pollination induced the concentration of actin bundles at the pollen attachment site, whereas self-pollination induced actin depolymerization; in addition, the actin-depolymerizing drug cytochalasin D significantly inhibited pollen germination during cross-pollination (Iwano et al, 2007). The pollen surface is covered with exine and a protein-and lipid-rich substance called the pollen coat.…”

Achievement of genetics in plant reproduction research: the past decade for the coming decade

“…In living pollen, visualization of mitochondria and plastids has been established (Matsushima et al, 2008;Tang et al, 2009), and precise monitoring of actin dynamics in papilla cells and Ca 2+ dynamics in pollen tubes is also successful in pollination studies (Iwano et al, 2007(Iwano et al, , 2009 …”