Periplasmic multilamellar membranous structures in Nicotiana tabacum L. pollen grains treated with Ni²âº or Cu²âº.

Essential trace elements Ni(2+) and Cu(2+) can block pollen germination without causing cell death. Mechanisms of this effect remain unclear. Using TEM, we studied the effects of Ni(2+) or Cu(2+) treatment on the ultrastructure of the aperture regions in tobacco pollen preparing to germinate in vitro, since in these zones, the main fluxes of water, ions, and metabolites cross the plasmalemma. Neither Ni(2+) nor Cu(2+) altered the cytoplasm ultrastructure, but both affected the reorganization of apertural periplasm during pollen activation. Numerous multilamellar membranous structures continuous with the plasma membrane could be seen in hydrated but not yet activated pollen. When the normal activation was completed, the structures disappeared and the plasmalemma became smooth. In the presence of 1 mM Ni(2+) or 100 µM Cu(2+), these structures preserved its original appearance. It is assumed to be the storage form for the membrane material, which is to provide an initial phase of the pollen tube growth. Ni(2+) and Cu(2+) affect the utilization of these membranes, thereby, blocking the pollen germination.

Ni(2+) effects on Nicotiana tabacum L. pollen germination and pollen tube growth.

To investigate the mechanisms of Ni(2+) effects on initiation and maintenance of polar cell growth, we used a well-studied model system-germination of angiosperm pollen grains. In liquid medium tobacco pollen grain forms a long tube, where the growth is restricted to the very tip. Ni(2+) did not prevent the formation of pollen tube initials, but inhibited their subsequent growth with IC(50) = 550 µM. 1 mM Ni(2+) completely blocked the polar growth, but all pollen grains remained viable, their respiration was slightly affected and ROS production did not increase. Addition of Ni(2+) after the onset of germination had a bidirectional effect on the tubes development: there was a considerable amount of extra-long tubes, which appeared to be rapidly growing, but the growth of many tubes was impaired. Studying the localization of possible targets of Ni(2+) influence, we found that they may occur both in the wall and in the cytoplasm, as confirmed by specific staining. Ni(2+) disturbed the segregation of transport vesicles in the tips of these tubes and significantly reduced the relative content of calcium in the aperture area of pollen grains, as measured by X-ray microanalysis. These factors are considered being critical for normal polar cell growth. Ni(2+) also causes the deposition of callose in the tips of the tube initials and the pollen tubes that had stopped their growth. We can assume that Ni(2+)-induced disruption of calcium homeostasis can lead to vesicle traffic impairment and abnormal callose deposition and, consequently, block the polar growth.