ROS in tobacco stigma exudate affect pollen proteome and provoke membrane hyperpolarization

1.AbstractROS are known to be accumulated in stigmas of different species and can possibly perform different functions in plant reproduction. Here we confirm the assumption that they affect pollen by altering ion transport through the plasma membrane; as a more deferred effect, pollen proteome is modified. We detected ROS in stigma exudate, found hyperpolarization in exudate-treated growing pollen tubes and used flow cytometry of pollen protoplasts to compare the effects of fresh exudate and exogenous H2O2 on pollen tube plasmalemma. Exudate causes plasmalemma hyperpolarization similar to the one provoked by H2O2, which is abolished by catalase treatment and ROS quencher MnTMPP. Inhibitory analysis indicates the participation of Ca2+- and K+-conducting channels in the observed hyperpolarization, linking obtained data with previous patch-clamp studies in vitro. For a deeper understanding of pollen response to ROS we analyzed proteome alterations in H2O2-treated pollen grains. We found 50 unique proteins and 20 differently accumulated proteins that are mainly involved in cell metabolism, energetics, protein synthesis and folding. Thus, pollen is getting ready for effective resource usage, construction of cellular components and rapid growth.HighlightsThe active substance in stigma exudate is H2O2H2O2 causes hyperpolarization mediated by the activation of cation channels.H2O2 affects pollen proteome; we found 50 unique proteins.

Nectar- and stigma exudate-specific expression of an acidic chitinase could partially protect certain apple cultivars against fire blight disease

Main conclusion Certain apple cultivars accumulate to high levels in their nectar and stigma exudate an acidic chitinase III protein that can protect against pathogens including fire blight disease causing Erwinia amylovora. Abstract To prevent microbial infections, flower nectars and stigma exudates contain various antimicrobial compounds. Erwinia amylovora, the causing bacterium of the devastating fire blight apple disease, is the model pathogen that multiplies in flower secretions and infects through the nectaries. Although Erwinia-resistant apples are not available, certain cultivars are tolerant. It was reported that in flower infection assay, the ‘Freedom’ cultivar was Erwinia tolerant, while the ‘Jonagold’ cultivar was susceptible. We hypothesized that differences in the nectar protein compositions lead to different susceptibility. Indeed, we found that an acidic chitinase III protein (Machi3-1) selectively accumulates to very high levels in the nectar and the stigma exudate of the ‘Freedom’ cultivar. We show that three different Machi3-1 alleles exist in apple cultivars and that only the 5B-Machi3-1 allele expresses the Machi3-1 protein in the nectar and the stigma exudate. We demonstrate that the 5B-Machi3-1 allele was introgressed from the Malus floribunda 821 clone into different apple cultivars including the ‘Freedom’. Our data suggest that MYB-binding site containing repeats of the 5B-Machi3-1 promoter is responsible for the strong nectar- and stigma exudate-specific expression. As we found that in vitro, the Machi3-1 protein impairs growth and biofilm formation of Erwinia at physiological concentration, we propose that the Machi3-1 protein could partially protect 5B-Machi3-1 allele containing cultivars against Erwinia by inhibiting the multiplication and biofilm formation of the pathogen in the stigma exudate and in the nectar.

Nectar- and stigma-specific expression of a chitinase could partially protect against fire blight in certain apples

To attract pollinators many angiosperms secrete stigma exudate and nectar in their flowers. As these nutritious fluids are ideal infection points for pathogens, both secretions contain various antimicrobial compounds. Erwinia amylovora, the causing bacterium of the devastating fire blight apple disease, is the model pathogen that multiplies in flower secretions and infects through the nectaries. Although Erwinia resistant apples are not available, certain cultivars are tolerant. It was reported that in stigma infection assay, the ‘Freedom’ cultivar was Erwinia tolerant while the ‘Jonagold’ was susceptible. We hypothesized that differences in the nectar protein compositions lead to different susceptibility. Indeed we found that an acidic chitinase III protein (Machi3-1) selectively accumulates in the nectar and stigma of the ‘Freedom’ cultivar. We demonstrate that MYB binding site containing repeats of the ‘Freedom’ Machi3-1 promoter are responsible for the strong nectar- and stigma-specific expression. As we found that in vitro the Machi3-1 protein impairs growth and biofilm formation of Erwinia at physiological concentration, we propose that the Machi3-1 contribute to the tolerance by inhibiting Erwinia multiplication in the stigma exudate and in the nectar. We show that the Machi3-1 allele was introgressed from Malus floribunda 821 into different apple cultivars including the ‘Freedom’.HighlightCertain apple cultivars accumulate to high levels in their nectar and stigma an acidic chitinase III protein that can protect against pathogens including fire blight disease causing Erwinia amylovora

Reproductive Function in the Mimosoid Legume Acacia retinodes: Ultrastructural and Cytochemical Characteristics of Stigma Receptivity

The stigma of Acacia retinodes is receptive from the moment the flower opens (female phase). Receptivity, assessed in terms of pod set, is highest at flower opening, and lowest the following day (male phase). Stigma receptivity is associated with the acquisition of a heterogeneous stigma exudate, the components of which are secreted sequentially during differentiation. Cytochemical probes have tentatively identified the principal components as unsaturated and saturated lipids, free fatty acids, flavonoid aglycones, carbohydrates, proteins and phenolic compounds. The onset of male phase in unpollinated stigmas is associated with a breakdown of the plasma membrane and organelle membranes of stigma cells, and subsequently a browning reaction.