Developmental and Structural Section
Ge, Lili , Gou, Xiaoping , Russell, Scott D. .
Dynamics of the angiosperm male germ unit during conveyance of sperm cells in Arabidopsis thaliana: Observation of GFP fluorescent cells in elongating pollen tubes.
Observations of living sperm cells in flowering plants are impeded by the near transparency of these cells in elongating pollen tubes. We therefore introduced a molecular gene construct consisting of sperm-specific promoter fused to a green fluorescent protein (GFP) gene. Transformed plants of Arabidopsis thaliana express the GFP construct in living sperm cells, which are strongly fluorescent. To observe sperm movement in tubes, pollen grains were germinated on 1% agar in 0.01% boric acid, 5 mM CaCl2, 5 mM KCl, 1 mM MgSO4, and 10% sucrose, at pH 7.5. Under these conditions, tubes reached >1 mm and continued vigorous elongation for the duration of observation (~6 hr). Sperm cells were tracked using epifluorescent, conventional confocal and spinning disc confocal microscopy. Intact sperm cells are quite variable in size and shape, but are typically elongated and spindle-shaped, bearing a projection at the end of the MGU that typically leads the cell during transport. The sperm cells and their slender projection are pleiomorphic and MGU appearance and position are quite dynamic. In vigorously elongating tubes, sperm cells may move by several micrometers per minute, varying on a second-by-second basis. Rather than being passively conveyed in the tube, they appear to be actively conveyed through a mechanism that requires the involvement of pollen tube cytoplasm. Directional movement of the cells appears to require the pollen-origin membrane around the MGU to be intact. Experimentally ablating the membranes enclosing these cells result in sperm cells becoming spherical and they lose their ability to track the movement of the other sperm cell. Although intact sperm cells undergo a net rate of transport that matches the elongation of the pollen tube, the velocity at any one time is quite variable. These observations establish the importance of the MGU as a requirement for success in double fertilization.
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1 - University of Oklahoma, Department of Botany and Microbiology, 770 Van Vleet Oval, Norman, OK, 73019, USA
2 - University of Oklahoma, Department of Botany and Microbiology, 770 Van Vleet Oval, Norman, Oklahoma, 73019, USA
male germ unit
Presentation Type: Oral Paper:Papers for Sections
Date: Tuesday, July 29th, 2008
Time: 10:45 AM