Micromanipulation of statoliths in gravity-sensing Chara rhizoids by optical tweezers

Abstract

Infrared laser traps (optical tweezers) were used to micromanipulate statoliths in gravity-sensing rhizoids of the green alga Chara vulgaris Vail. We were able to hold and move statoliths with high accuracy and to observe directly the effects of statolith position on cell growth in horizontally positioned rhizoids. The first step in gravitropism, namely the physical action of gravity on statoliths, can be simulated by optical tweezers. The direct laser microirradiation of the rhizoid apex did not cause any visible damage to the cells. Through lateral positioning of statoliths a differential growth of the opposite flank of the cell wall could be induced, corresponding to bending growth in gravitropism. The acropetal displacement of the statolith complex into the extreme apex of the rhizoid caused a temporary decrease in cell growth rate. The rhizoids regained normal growth after remigration of the statoliths to their initial position 10-30 micrometers basal to the rhizoid apex. During basipetal displacement of statoliths, cell growth continued and the statoliths remigrated towards the rhizoid tip after release from the optical trap. The resistance to statolith displacement increased towards the nucleus. The basipetal displacement of the whole complex of statoliths for a long distance (>100 micrometers) caused an increase in cell diameter and a subsequent regaining of normal growth after the statoliths reappeared in the rhizoid apex. We conclude that the statolith displacement interferes with the mechanism of tip growth, i.e. with the transport of Golgi vesicles, either directly by mechanically blocking their flow and/or, indirectly, by disturbing the actomyosin system. In the presence of the actin inhibitor cytochalasin B the optical forces required for acropetal and basipetal displacement of statoliths were significantly reduced to a similar level. The lateral displacement of statoliths was not changed by cytochalasin B. The results indicate: (i) the viscous resistance to optical displacement of statoliths depend mainly on actin, (ii) the lateral displacement of statoliths is not impeded by actin filaments, (iii) the axially directed actin-mediated forces against optical displacement of statoliths (for a distance of 10 micrometers) are stronger in the basipetal than in the acropetal direction, (iv) the forces acting on single statoliths by axially oriented actin filaments are estimated to be in the range of 11-110 pN for acropetal and of 18-180 pN for basipetal statolith displacements.

Optical trapping and manipulation of neutral particles using lasers

“…The three computer-controlled tweezers traps used are an application of a multiple scanning trap system developed by Visscher et al (139). Greulich et al have used tweezers to simulate the effect of gravity on the growing tips of algal cells (140). Dragging the statoliths or gravity sensors of the cell to one side can induce the cell to reorient its growth in that direction.…”

Gravitropism in tip-growing cells

“…(ii) Leitz et al (1995) initiated bending of rhizoids by using optical tweezers to translocate statoliths laterally to one¯ank of the rhizoids. The laser output power necessary for moving statoliths in the basal direction was higher than that for moving statoliths in the apical direction.…”

Statolith Sedimentation Kinetics and Force Transduction to the Cortical Endoplasmic Reticulum in Gravity-SensingArabidopsisColumella Cells

“…In this study, we applied differential interference contrast (DIC) microscopy, an imaging technique that generates minimal photodamage during real-time observations (Danuser et al, 2000), and laser tweezers, a noninvasive technique to apply controlled forces inside living cells (Ashkin and Dziedzic, 1989;Leitz et al, 1995), to analyze the movements of statoliths in relation to the cortical ER interface in columella cells of intact Arabidopsis seedlings. In addition, we applied high-pressure freezing and electron tomography to characterize the statolith-induced changes in ER membrane structure with nanometer resolution in Arabidopsis, Nicotiana tabacum, and Medicago sativa.…”