Analysis of Statoliths Displacement in Chara Rhizoids for Validating the Microgravity-Simulation Quality of Clinorotation Modes

Abstract

In single-celled rhizoids of the green algae Chara, positively gravitropic growth is governed by statoliths kept in a dynamically stable position 10-25 μm above the cell tip by a complex interaction of gravity and actomyosin forces. Any deviation of the tube-like cells from the tip-downward orientation causes statoliths to sediment onto the gravisensitive subapical cell flank which initiates a gravitropic curvature response. Microgravity experiments have shown that abolishing the net tip-directed gravity force results in an actomyosin-mediated axial displacement of statoliths away from the cell tip. The present study was performed to critically assess the quality of microgravity simulation provided by different operational modes of a Random Positioning Machine (RPM) running with one axis (2D mode) or two axes (3D mode) and different rotational speeds (2D), speed ranges and directions (3D). The effects of 2D and 3D rotation were compared with data from experiments in real microgravity conditions (MAXUS sounding rocket missions). Rotational speeds in the range of 60-85 rpm in 2D and 3D modes resulted in a similar kinetics of statolith displacement as compared to real microgravity data, while slower clinorotation (2-11 rpm) caused a reduced axial displacement and a more dispersed arrangement of statoliths closer to the cell tip. Increasing the complexity of rotation by adding a second rotation axis in case of 3D clinorotation did not increase the quality of microgravity simulation, however, increased side effects such as the level of vibrations resulting in a more dispersed arrangement of statoliths. In conclusion, fast 2D clinorotation provides the most appropriate microgravity simulation for investigating the graviperception mechanism in Chara rhizoids, whereas slower clinorotation speeds and rotating samples around two axes do not improve the quality of microgravity simulation.

Statoliths displacement in root statocytes in real and simulated microgravity

“…Organs and cells, which deflect the axis for some distance, will undergo centrifugal forces. A comparison of results from flight experiments and clinorotation with unicellular and multicellular organisms and tissue cultures showed that the fast clinostat is a valuable tool for evaluating an organism's sensitivity to altered gravity [8,9,37]. The absolute magnitude of the vector is incidentally preserved, consequently, the environmental properties are not changed.…”

“…For instance, the certain similarity of data on activation or inhibition of gene expression and protein synthesis associated with cell wall remodeling detected both in A. thaliana seedlings grown in real microgravity [38][39][40] and under 3-D clinorotation [41], as well as loosening and thinning of cell walls in microgravity and under clinostation can be explained in the light of biochemical data on quantitative and qualitative changes in the composition of poly-and monosaccharides [42,43]. A fast-rotating 2-D clinostat is also considered to be an efficient tool for studying the plant responses to simulated microgravity [8,15]. Changes in gene expression in roots of 5-day old A. thaliana seedlings as well as in the location of amyloplasts-statoliths in cap statocytes after 3 min and 6 min of fast 2-D clinirotation were described [15].…”

“…To simulate biological effects of microgravity in space flight, various ground-based facilities -slow (1-10 rpm) and fast (50-120 rpm) 2-D clinostats, Random Positioning Machine, Free Fall Machine, Rotating Wall Vessels and magnetic levitation -are widely used [1][2][3][4][5][6][7] as experiments in "real μg are scarce, costly and time-consuming" [8]. Capacities and limitations of various devices for accurate and reliable simulations of microgravity conditions comparable to real microgravity in space are constantly discussed [6,[9][10][11][12][13][14][15][16].…”

“…Recent comparative studies of the quality of microgravity simulation provided by different operational modes have shown the greatest suitability of fast 2-D clinorotation for investigating the gravi-perception mechanism in Chara rhizoids compared with slower 2-D and 3-D clinorotation and rotation of samples around two axes [8]. In the given paper, we compare the position of amyloplasts-statoliths in root cap statocytes of higher plants at 1 g, slow and fast 2-D clinorotation, real microgravity in space flight, vibration and acceleration in the spacecraft launch mode.…”

Guanylyl Cyclase-cGMP Signaling Pathway in Melanocytes: Differential Effects of Altered Gravity in Non-Metastatic and Metastatic Cells

“…It is therefore recommended to perform comparative studies due to the different working principles of the RPM and the clinostat. Furthermore, the quality of simulation should be validated under real microgravity (e.g., in space) [72].…”