News & Announcements
(July '07)
Our lab will present 5 posters in the upcoming Plant Biology2007, annual American Society of Plant Biologists conference in Chicago (July 7- 11, 2007).
Click on poster #s below to access abstracts in Plant Biology2007 website:
1098, 1311, 1907, 2259, 2282
The abstracts of the five poster presentations are:
1. Poster #: 1098
Qin, Yuan [1], Palanivelu, Ravishankar [2].
Identifying the pistil gene network that regulates final stages of pollen tube guidance.
A pollen tube navigates past different pistil cells before it reaches the egg to effect fertilization and disruption in pollen tube guidance results in no seed formation. Pistil cells provide nutritional and navigation cues to guide pollen tubes to ovules, yet only a few have been identified. We are using arabidopsis actin-related protein 6 (ARP6) to isolate signals from female sporophytic cells that mediate the final stages of pollen tube growth to ovules. ARP6 is a component of chromatin remodeling complex involved in regulating gene expression. arp6 mutant plants are characterized by defects in pistil tissues causing low seed set. To explore the basis of low seed set, Lat52:GUS pollen was grown on WT or arp6 pistils and stained for Gus activity. In the arp6 pistils, although pollen tube germination and growth were comparable to WT, only 54.28% of ovules were targeted by pollen tubes compared to WT (86.90%). In order to pinpoint which arp6 pistil tissue is defective, we performed in vitro pollen tube guidance assay involving several different combinations of WT and arp6 pistil and ovules. When Lat52:GFP pollen tubes approached WT ovules after traveling through WT or arp6 pistils, ~50% of the ovules were targeted. However, if the LAT52:GFP pollen tubes grew through WT or arp6 pistils and approached arp6 ovules, only 35% of the ovules were targeted suggesting that the arp6 ovule defects resulted in low seed set. To identify the ovule gene network associated with final stages of pollen tube guidance, we compared gene expression changes in arp6 and WT ovules by microarray analysis. Progress on the microarray experiments and characterization of these genes will be reported.
Keywords:
pollen tube guidance
Arabidopsis
arp6.
Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P28035
Abstract ID:1098
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2. Poster #: 1311
Palanivelu, Ravishankar [1], Drews, Gary [2].
Synergid Cell Death in Arabidopsis is Triggered Following Direct Interaction With the Pollen Tube.
During angiosperm reproduction, one of the two synergid cells within the female gametophyte undergoes cell death prior to fertilization. Synergid degeneration is an essential step because the pollen tube enters the female gametophyte by growing into the synergid cell that undergoes cell death and releases its two sperm cells within the degenerating synergid cytoplasm to effect double fertilization. In Arabidopsis and many other species, synergid cell death is dependent upon pollination. However, the mechanism by which the pollen tube causes synergid cell death is not understood.
As a first step toward understanding this mechanism, we defined the temporal relationship between pollen tube arrival at the female gametophyte and synergid cell death by performing live-imaging of pollen tube growth and synergid cell death using an in vitro pollen tube guidance assay. To observe pollen tube growth, pollen tubes were marked with DsRed driven by the pollen-specific LAT52 promoter. To observe synergid degeneration, we specifically labeled the synergid cells with green fluorescent protein (GFP) expressed from the MYB98 promoter and used loss of the synergid GFP signal as an indicator of synergid degeneration. Using real-time observation of these two events in vitro, we demonstrate that synergid cell death initiates after the pollen tube arrives at the female gametophyte but before pollen tube discharge. Our results support a model in which the pollen tube induces a physiological cell death program following direct interaction with the synergid cell.
Keywords:
pollen tube
synergid
Programmed Cell Death
degeneration
female gametophyte
embryosac
guidance.
Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P28038
Abstract ID:1311
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3. Poster #: 1907
Tsukamoto, Tatsuya [1], Drews, Gary [2], Palanivelu, Ravishankar [3].
Characterization of a female gametophytic Arabidopsis mutant defective in pollen tube guidance.
A pollen tube encounters many interactions with female gametophytic cells on its journey to female gametophyte such as attraction, repulsion and adhesion. This journey begins with pollen tube growing between the walls of stigma cells, traveling through the extracellular matrix of the transmitting tract, and finally arriving at the ovary, where it migrates up the funiculus, and enter the micropyle of an ovule to deliver the two sperm cells-one fertilizes an egg and other the central cell. Thus a pollen tube navigates past several different female cells before it reaches the egg. Signals that mediate the migration of a pollen tube to an ovule remain poorly characterized. A female gametophytic mutant, fem137 was obtained by screening for plants with reduced seed set from a T-DNA insertion collection. Siliques in fem137/+ plants contain significantly reduced number of seeds (30-40 seeds per silique) compared to wild-type (~ 60 seeds per silique). To explore the basis of reduced seed set, we pollinated fem137/+ pistils with LAT52:GUS pollen and stained for GUS activity. In the fem137/+ pistils, although pollen tube germination and growth in transmitting tract was normal, nearly half of the ovules were not targeted by the pollen tube and remained unfertilized. Since the kanamycin marker on the T-DNA does not co-segregate with the reduced seed set phenotype, we are using map-based cloning procedure to identify the gene(s) defined by fem137 mutation. Progress on the cloning and characterization of fem137 will be presented.
Keywords:
Arabidopsis
Female gametophytic mutant
fem137
pollen tube guidance.
Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P28051
Abstract ID:1907
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4. Poster #: 2259
Huang, Yiding [1], Palanivelu, Ravishankar [2].
Female gametophytic mutant impaired in final stages of pollen tube guidance.
Pollen tube growth and guidance initiates after pollen grain hydrates on the papillar cells of stigma. Subsequently, pollen tube is guided along the transmission tissue, the funiculus and then into the ovule micropyle. After entering the ovule, pollen tube terminates its journey within the synergid cell. Seeds are not produced if any aspect of this pollen tube guidance process is disrupted. Although some of the factors affecting pollen tube growth and guidance have been identified, many aspects of pollen tube guidance still remain unclear. Arabidopsis female gametophytic mutants defective in pollen tube growth and guidance are ideal tools to unravel the mechanisms that regulate pollen tube guidance to embryo sacs. For this purpose, we screened Arabidopsis plants with reduced seed containing siliques from among the SALK TDNA insertion collection and identified a reduced fertile mutant (ref1, average seed number 22.54±5.75 compared to 56.17±3.37 in wild type). Based on the progeny analyses and reciprocal crosses to wild type plants, it was found that ref1 caused female gameotphytic defects. To identify the pollen tube growth stage that is defective in ref1, we pollinated mutant pistils with Lat52:GUS pollen and monitored the GUS activity. These experiments demonstrated that pollen tube germination and growth in mutant stigma, style and ovary were comparable to pollen tube behavior on a wild type pistil. However, even wild type pollen tubes exhibited abnormal and excessive growth after entering the mutant ovules. Real-time observations of pollen tube interactions with ref1 mutant ovules in an in vitro guidance assay also revealed that fertilized ref1 ovules were defective in deflecting additional pollen tubes. Progress on microscopy analyses of ref1 mutant phenotypes and cloning the gene disrupted in ref1 will be reported.
Keywords:
Female gametophytic mutant
pollen tube guidance.
Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P28056
Abstract ID:2259
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5. Poster #: 2282
Jackson, Jamie [1], Palanivelu, Ravishankar [1].
An Arabidopsis mutant defective in pollen tube repulsion.
After pollen lands on the surface of the pistil, it extends a long polar process that carries sperm to an ovule. Typically, only one pollen tube successfully enters the ovule micropyle and terminates its journey within a synergid cell by bursting to release sperm cells. Since ovules are penetrated by only one pollen tube, it has been hypothesized that additional tubes are actively repulsed by a targeted ovule. Previously, it has been shown that Arabdiopsis thaliana in vitro guidance assay recapitulates pollen tube repulsion near a targeted ovule. By performing real-time observations of pollen tube interactions with a targeted ovule, we first explored the divergence of repulsion mechanism that prevents multiple tubes from entering a targeted ovule. Characterization of pollen tube repulsion was carried out using repulsion data from two close species of A. thaliana; A. arenosa and Olimarabidopsis pumila. Results from these experiments demonstrated that even if targeted by A. thaliana pollen tubes, A. arenosa (25 instances, n=20 assays) and Olimarabidopsis pumila ovules (10 instances, n=20 assays) repel additional tubes that approach them. These results suggest that pollen tube repulsion is mediated by a signaling mechanism that does not evolve rapidly. Secondly, we used the in vitro guidance assay to characterize the pollen tube repulsion defects in a newly isolated mutant, ref1 (reduced fertility defect 1). Mutant ovules that have been already targeted by pollen tubes are markedly defective in repelling additional tubes that approach them (5 out of 12 instances, ~42%; n=20 assays) resulting in multiple tube entry into the micropyle. This percentage is significantly different from the behavior of wild type ovules in the in vitro assay; in every instance, wild type targeted ovules repelled additional tubes that approached them (0 out of 44. 0%; n=20 assays). Lack of repulsion in ref1 may point to decreased production of repellants upon ovule targeting.
Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P34017
Abstract ID:2282
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