Pollen Tubes Fail to Target Ovule in the Absence of Two Cation/proton Exchangers in Arabidopsis

Title of Document: POLLEN TUBES FAIL TO TARGET OVULE IN THE ABSENCE OF TWO CATION/PROTON EXCHANGERS IN ARABIDOPSIS Yongxian Lu Doctor of Philosophy, 2010 Directed By: Professor Heven Sze Department of Cell Biology & Molecular Genetics Flowering plant reproduction requires precise delivery of the sperm cells to the ovule by a pollen tube. Guiding signals from female cells are being identified, though how pollen senses and responds to those cues are largely unknown. Here I provide genetic evidence that two predicted cation/proton exchangers expressed in Arabidopsis pollen play essential roles in pollen targeting of ovules. Male fertility was unchanged in single chx21 or chx23 mutant pollen; however, male-specific gene transmission was blocked in chx21chx23 double mutant. Wild-type pistil provided with a limited amount of pollen containing a mixture of single and double mutant produced ~60% less seeds compared to that produced with chx23 single mutant pollen, indicating that chx21chx23 pollen is infertile. The double mutant pollen, visualized by a pollen-specific promoter-driven GUS activity, germinated and extended a tube down the transmitting tract, but the tube failed to turn and target an ovule. Unlike wild-type pollen that targeted isolated ovules in a semi-in vivo assay, tube guidance in chx21chx23 pollen was compromised. As a first step to understand the cellular and molecular bases of tube guidance, membrane localization and activity of CHX23 was determined. GFP-tagged CHX23 was localized to endomembranes, predominantly endoplasmic reticulum (ER), in elongating pollen tubes. Furthermore, expression of CHX23 in E. coli resulted in enhanced K + accumulation at alkaline pH, suggesting a role for CHX23 in K + acquisition and pH homeostasis. Based on these studies and observations by others that ER oscillates and enters the apex, a simple model is proposed: Modification of localized pH by CHX21 or CHX23 enables pollen tube to sense female signals and respond by shifting directional growth at the funiculus and micropyle to target pollen tip growth towards the ovule. POLLEN TUBES FAIL TO TARGET OVULE IN THE ABSENCE OF TWO CATION/PROTON EXCHANGERS IN ARABIDOPSIS