Detection and localization of pectin methylesterase isoforms in pollen tubes of Nicotiana tabacum L.

Pectin methylesterases (PMEs) were detected in tobacco (Nicotiana tabacum) pollen tubes grown in vitro. Seven PME isoforms exhibiting a wide isoelectric-point (pI) range (5.3-9.1) were found in crude extracts of pollen tubes. These isoforms were mainly retrieved in supernatants after low- and high-speed separation of the crude extract. Two isoforms, with pIs 5.5 and 7.3 and molecular weight about 158 kDa, were detected by immunoblotting with anti-flax PME antiserum. Localization of pectins and PME isoforms in pollen tubes was investigated by immunogold labelling with JIM5 monoclonal antibodies and anti-flax PME antiserum, respectively. In germinated pollen grains, two PME isoforms were mainly detected in the exine, Golgi apparatus and secretory vesicles. In pollen tubes the same two PME isoforms were distributed along the outer face of the plasma membrane in the vicinity of the inner layer of the cell wall, in the Golgi and around secretory vesicles. In pollen grains, PME isoforms were, in some cases, mixed with acidic pectins in proximity to the outer surface of the plasma membrane. In pollen tubes the presence of PMEs inside secretory vesicles carrying esterified pectins supports the hypothesis that, during pollen tube growth, PMEs could be transferred by secretory vesicles in a precursor form and be activated at the tip where exocytosis takes place.