Calcium gradients in conifer pollen tubes; dynamic properties differ from those seen in angiosperms.

Pollen tubes are an established model system for examining polarized cell growth. The focus here is on pollen tubes of the conifer Norway spruce (Picea abies, Pinaceae); examining the relationship between cytosolic free Ca2+, tip elongation, and intracellular motility. Conifer pollen tubes show important differences from their angiosperm counterparts; they grow more slowly and their organelles move in an unusual fountain pattern, as opposed to reverse fountain, in the tip. Ratiometric ion imaging of growing pollen tubes, microinjected with fura-2-dextran, reveals a tip-focused [Ca2+]i gradient extending from 450 nM at the extreme apex to 225 nM at the base of the tip clear zone. Injection of 5,5' dibromo-BAPTA does not dissipate the apical gradient, but stops cell elongation and uniquely causes rapid, transient increases of apical free Ca2+. The [Ca2+]i gradient is, however, dissipated by reversible perfusion of extracellular caffeine. When the basal cytosolic free Ca2+ concentration falls below 150 nM, again a large increase in apical [Ca2+]i occurs. An external source of calcium is not required for germination but significantly enhances elongation. However, both germination and elongation are significantly inhibited by the inclusion of calcium channels blockers, including lanthanum, gadolinium, or verapamil. Modulation of intracellular calcium also affects organelle position and motility. Extracellular perfusion of lanthanides reversibly depletes the apical [Ca2+]i gradient, altering organelle positioning in the tip. Later, during recovery from lanthanide perfusion, organelle motility switches direction to a reverse fountain. When taken together these data show a unique interplay in Picea abies pollen tubes between intracellular calcium and the motile processes controlling cellular organization.

Effect of extracellular calcium, pH and borate on growth oscillations in Lilium formosanum pollen tubes.

Calcium ions (Ca(2+)), protons (H(+)), and borate (B(OH)(4)(-)) are essential ions in the control of tip growth of pollen tubes. All three ions may interact with pectins, a major component of the expanding pollen tube cell wall. Ca(2+ )is thought to bind acidic residues, and cross-link adjacent pectin chains, thereby strengthening the cell wall. Protons are loosening agents; in pollen tube walls they may act through the enzyme pectin methylesterase (PME), and either reduce demethylation or stimulate hydrolysis of pectin. Finally, borate cross-links monomers of rhamnogalacturonan II (RG-II), and thus stiffens the cell wall. It is demonstrated here that changing the extracellular concentrations of Ca(2+), H(+) and borate affect not only the average growth rate of lily pollen tubes, but also influence the period of growth rate oscillations. The most dramatic effects are observed with increasing concentrations of Ca(2+) and borate, both of which markedly reduce the rate of growth of oscillating pollen tubes. Protons are less active, except at pH 7.0 where growth is inhibited. It is noteworthy, especially with borate, that the faster growing tubes exhibit the shorter periods of oscillation. The results are consistent with the idea that binding of Ca(2+) and borate to the cell wall may act at a similar level to alter the mechanical properties of the apical cell wall, with optimal concentrations being high enough to impart sufficient rigidity to the wall so as to prevent bursting in the face of cell turgor, but low enough to allow the wall to stretch quickly during periods of accelerating growth.

Control of pollen tube growth: role of ion gradients and fluxes.

Pollen tube growth attracts our attention as a model system for studying cell elongation in plants. The process is fast, it is confined to the tip of the tube, and it is crucial for sexual reproduction in plants. In the enclosed review we focus on the control of pollen tube growth, giving special attention to the role of ions, especially calcium and protons. During the last decade technical advances have made it possible to detect localized intracellular gradients, and extracellular fluxes of calcium and protons in the apical domain. Other ions, notably potassium and chloride, are also receiving attention. An important development has been the realization that pollen tube growth oscillates in rate; in addition, the ion gradients and fluxes oscillate in magnitude. Although all the ionic oscillations show the same period as that of the growth rate, with the exception of extracellular chloride efflux, they are not in phase with growth. Considerable effort is devoted to the elucidation of these different phase relationships, with the view that a hierarchical order may provide clues about those events that are primary vs. secondary in growth control. Attention is also given to the targets for the ions, for example, the secretory system, the cytoskeleton, the cell wall, in an attempt to provide a global understanding of pollen tube growth. Contents Summary 539 I. Introduction 540 II. Ion gradients and flux patterns 541 III. Oscillations 544 IV. The need for a Ca2+ store 547 V. Intracellular targets for Ion activity 549 VI. Extracellular targets for ions: the cell wall 552 VII. Ions in navigation 554 VIII. Role of ions in self-incompatibility 555 IX. The plasma membrane; site of global coordination and control 556 X. A model for pollen tube growth 557 IX. Conclusions 558 Acknowledgements 559 References 559.

Involvement of extracellular calcium influx in the self-incompatibility response of Papaver rhoeas.

We have previously demonstrated that increases in cytosolic free Ca2+ are triggered by the self-incompatibility (SI) response in incompatible Papaver rhoeas (the field poppy) pollen. However, one key question that has not been answered is whether extracellular Ca2+ may be involved. To address this question, we have used an ion-selective vibrating probe to measure changes in extracellular Ca2+ fluxes around poppy pollen tubes. Our data reveal several findings. First, we confirm that there is an oscillating Ca2+ influx directed at the apex of the pollen tube; we also provide evidence that Ca2+ influx also occurs at the shanks of pollen tubes. Second, upon challenge with self-incompatibility (S) proteins, there is a stimulation of Ca2+ influx along the shank of incompatible pollen tubes, approximately 50 microm behind the pollen tube tip. This demonstration of SI-induced Ca2+ influx suggests a role for influx of extracellular Ca2+ in the SI response.