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Magnitude and Direction of Vesicle Dynamics in Growing Pollen Tubes Using Spatiotemporal Image Correlation Spectroscopy and Fluorescence Recovery after Photobleaching^1,[W],[OA]

Institut de recherche en biologie végétale, Département de sciences biologiques, Université de Montréal, Montreal, Quebec, Canada H1X 2B2 (J.B., A.G.); Department of Physics, McGill University, Montreal, Quebec, Canada H3A 2T8 (B.V., J.K., P.W.W.); Biology Department, University of Massachusetts, Amherst, Massachusetts 01003 (P.K.H.); and Department of Chemistry, McGill University, Montreal, Quebec, Canada H3A 2K6 (P.W.W.)

The delivery of cell wall material and membrane to growing plant cell surfaces requires the spatial and temporal coordination of secretory vesicle trafficking. Given the small size of vesicles, their dynamics is difficult to quantify. To quantitatively analyze vesicle dynamics in growing pollen tubes labeled with the styryl dye FM1-43, we applied spatiotemporal correlation spectroscopy on time-lapse series obtained with high-speed confocal laser scanning microscopy recordings. The resulting vector maps revealed that vesicles migrate toward the apex in the cell cortex and that they accumulate in an annulus-shaped region adjacent to the extreme tip and then turn back to flow rearward in the center of the tube. Fluorescence recovery after photobleaching confirmed vesicle accumulation in the shoulder of the apex, and it revealed that the extreme apex never recovers full fluorescence intensity. This is consistent with endocytotic activity occurring in this region. Fluorescence recovery after photobleaching analysis also allowed us to measure the turnover rate of the apical vesicle population, which was significantly more rapid than the theoretical rate computed based on requirements for new cell wall material. This may indicate that a significant portion of the vesicles delivered to the apex does not succeed in contacting the plasma membrane for delivery of their contents. Therefore, we propose that more than one passage into the apex may be needed for many vesicles before they fuse to the plasma membrane and deliver their contents.

The authors responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) are: Anja Geitmann (anja.geitmann{at}umontreal.ca) and Paul W. Wiseman (paul.wiseman{at}mcgill.ca; STICS software).

^[W] The online version of this article contains Web-only data.

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www.plantphysiol.org/cgi/doi/10.1104/pp.108.120212

^* Corresponding author; e-mail anja.geitmann{at}umontreal.ca.

Received April 1, 2008; accepted May 19, 2008; published May 28, 2008.

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