The guidance signals that drive pollen tube navigation inside the pistil and micropyle targeting are still, to a great extent, unknown. Previous studies in vitro showed that nitric oxide （NO） works as a negative che...The guidance signals that drive pollen tube navigation inside the pistil and micropyle targeting are still, to a great extent, unknown. Previous studies in vitro showed that nitric oxide （NO） works as a negative chemotropic cue for pollen tube growth in lily （Lilium Iongiflorum）. Furthermore, Arabidopsis thaliana Atnosl mutant plants, which show de- fective NO production, have reduced fertility. Here, we focus in the role of NO in the process of pollen-pistil communication, using Arabidopsis in-vivo and lily semi-vivo assays. Cross-pollination between wild-type and Atnosl plants shows that the mutation affects the pistil tissues in a way that is compatible with abnormal pollen tube guidance. Moreover, DAF- 2DA staining for NO in kanadi floral mutants showed the presence of NO in an asymmetric restricted area around the micropyle. The pollen-pistil interaction transcriptome indicates a time-course-specific modulation of transcripts of AtNOS1 and two Nitrate Reductases （nrl and nr2）, which collectively are thought to trigger a putative NO signaling pathway. Semivivo assays with isolated ovules and lily pollen further showed that NO is necessary for micropyle targeting to occur. This evidence is supported by CPTIO treatment with subsequent formation of balloon tips in pollen tubes facing ovules. Activation of calcium influx in pollen tubes partially rescued normal pollen tube morphology, suggesting that this pathway is also dependent on Ca^2＋ signaling. A role of NO in modulating Ca^2＋ signaling was further substantiated by direct imaging the cytosolic free Ca^2＋ concentration during NO-induced re-orientation, where two peaks of Ca^2＋ occur--one during the slowdown/stop response, the second during re-orientation and growth resumption. Taken together, these results provide evidence for the participation of NO signaling events during pollen-pistil interaction. Of special relevance, NO seems to directly affect the targeting of pollen tubes to the ovule＇s micropyle by modulating the action of its diffusible factors.展开更多
文摘The guidance signals that drive pollen tube navigation inside the pistil and micropyle targeting are still, to a great extent, unknown. Previous studies in vitro showed that nitric oxide （NO） works as a negative chemotropic cue for pollen tube growth in lily （Lilium Iongiflorum）. Furthermore, Arabidopsis thaliana Atnosl mutant plants, which show de- fective NO production, have reduced fertility. Here, we focus in the role of NO in the process of pollen-pistil communication, using Arabidopsis in-vivo and lily semi-vivo assays. Cross-pollination between wild-type and Atnosl plants shows that the mutation affects the pistil tissues in a way that is compatible with abnormal pollen tube guidance. Moreover, DAF- 2DA staining for NO in kanadi floral mutants showed the presence of NO in an asymmetric restricted area around the micropyle. The pollen-pistil interaction transcriptome indicates a time-course-specific modulation of transcripts of AtNOS1 and two Nitrate Reductases （nrl and nr2）, which collectively are thought to trigger a putative NO signaling pathway. Semivivo assays with isolated ovules and lily pollen further showed that NO is necessary for micropyle targeting to occur. This evidence is supported by CPTIO treatment with subsequent formation of balloon tips in pollen tubes facing ovules. Activation of calcium influx in pollen tubes partially rescued normal pollen tube morphology, suggesting that this pathway is also dependent on Ca^2＋ signaling. A role of NO in modulating Ca^2＋ signaling was further substantiated by direct imaging the cytosolic free Ca^2＋ concentration during NO-induced re-orientation, where two peaks of Ca^2＋ occur--one during the slowdown/stop response, the second during re-orientation and growth resumption. Taken together, these results provide evidence for the participation of NO signaling events during pollen-pistil interaction. Of special relevance, NO seems to directly affect the targeting of pollen tubes to the ovule＇s micropyle by modulating the action of its diffusible factors.
