Abstract
Pollen-pistil interactions are critical early events regulating pollination and fertilization. Self-incompatibility (SI) is an important mechanism to prevent self-fertilization and inbreeding in higher plants. Although data implicate the involvement of reactive oxygen species (ROS) and nitric oxide (NO) in pollen-pistil interactions and the regulation of pollen tube growth, there has been a lack of studies investigating ROS and NO signaling in pollen tubes in response to defined, physiologically relevant stimuli. We have used live-cell imaging to visualize ROS and NO in growing Papaver rhoeas pollen tubes using chloromethyl-2'7'-dichlorodihydrofluorescein diacetate acetyl ester and 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate and demonstrate that SI induces relatively rapid and transient increases in ROS and NO, with each showing a distinctive "signature" within incompatible pollen tubes. Investigating how these signals integrate with the SI responses, we show that Ca(2+) increases are upstream of ROS and NO. As ROS/NO scavengers alleviated both the formation of SI-induced actin punctate foci and also the activation of a DEVDase/caspase-3-like activity, this demonstrates that ROS and NO act upstream of these key SI markers and suggests that they signal to these SI events. These data represent, to our knowledge, the first steps in understanding ROS/NO signaling triggered by this receptor-ligand interaction in pollen tubes.
Highlights
Pollen-pistil interactions in flowering plants are involved in pivotal events regulating pollination and fertilization
We have used live-cell imaging to demonstrate that induction of SI by addition of a recombinant protein, PrsS to incompatible Papaver pollen, stimulates transient increases in both reactive oxygen species (ROS) and nitric oxide (NO) in response to SI in incompatible pollen
These data describe a physiologically relevant, defined protein ligand stimulus-induced ROS/NO response in pollen tubes, and implicate an involvement for both ROS and NO in the SI-mediated PCD-signaling network. They represent the first steps in investigating ROS/NO signaling triggered by receptor-ligand interactions in pollen tubes
Summary
Pollen-pistil interactions in flowering plants are involved in pivotal events regulating pollination and fertilization. An important mechanism that operates during pollination to prevent inbreeding and its consequential debilitating effects is Self-Incompatibility (SI). Three distinct SI systems have been identified to date at a molecular level, which suggests that SI has evolved independently several times; see (Takayama and Isogai, 2005; FranklinTong, 2008) for recent reviews. These systems use a variety of mechanisms to prevent selffertilization. Self-fertilization is prevented by use of a specific recognition system, which results in “self” pollen (i.e. incompatible pollen) being rejected at some point in the pollination process, while compatible pollen is allowed to grow freely. When pollen S- and pistil S-haplotypes match, this creates an incompatible combination and incompatible pollen is rejected
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