The effect of exposure to microgravity on the development and structural organisation of plant protoplasts flown on Biokosmos 9

Abstract

Preparatory experiments for the IML-1 (International Microgravity Laboratory) mission to be flown on the Space Shuttle in January, 1992, were performed on a 14 day flight on Biokosmos 9 (Kosmos 2044) in September 1989. The purpose of the experiment was to study the effect of weightlessness on protoplast regeneration. Problems with late access to the space vehicle meant that the newly isolated protoplasts from hypocotyl cells of rapeseed (Brassica napus L. cv Niklas) and suspension cultures of carrot (Daucus carota L, cv Nobo) had to be stored at 4 degrees C for 36 h prior to the launch of the biosatellite, in order to delay cell wall regeneration until the samples were in orbit. In the flight samples and the ground controls, a portion of the total number of protoplasts regenerated cell walls. The growth of flight rapeseed cells was only 56% compared to the ground control; the respective growth of carrot cells in orbit was 82% of the ground control. Analysis demonstrated that the peroxidase activity and the amount of protein was lower in the flight samples than in the ground controls. The number of different isoenzymes was also decreased in the flight samples. A 54% decrease in the production of cellulose was found in rapeseed, and a 71% decrease in carrot. Hemicellulose production was also decreased in the flight samples compared to the ground controls. Ultrastructural analysis of the cell aggregates from the protoplasts cultured in orbit, demonstrated that hydrolysis and disappearance of reserve starch occurred in the flight cell plastids. The mitochondria were more varied in appearance in the flight samples than in the ground control cells. An increased frequency of the occurrence of folds formed by the plasmalemma together with an increase in the degree of complexity of these folds was also observed. Fluorescence analysis showed a decrease of the calcium content in cell cultures under space flight compared to the ground controls. One general effect of the stay onboard the space vehicle was a retardation of the regeneration processes. Callus cultures obtained from the flight samples grew very slowly compared to callus regenerated from the ground controls, and two years after the Biokosmos 9 flight there appears to be no further growth in the samples exposed to microgravity. Callus cultures from the ground controls, however, continue to grow well. A simulation experiment for IML-l performed in January 1990 at ESTEC (European Space Technology Center), The Netherlands, has resulted in regenerated plants. These observations are discussed and compared to the results obtained on Biokosmos 9.

Simulated weightlessness and hyper‐g results in opposite effects on the regeneration of the cortical microtubule array in protoplasts from Brassica napus hypocotyls

“…Narrow polyethylene bags (2 mm in diameter) were used for the cultivation of the protoplasts and mounted in the mid‐axis of the horizontal clinostat. The polyethylene bags are made of the same material as the bags used in space experiments performed by our group (see Rasmussen et al 1992). The protoplasts were cultivated on the clinostat in the dark (25°C) for 5, 24, 48, 72 or 96 h. Exposure of the protoplasts to hyper ‐g (7–10 g ) was done by using a special centrifuge (A. Johnsson, unpublished data).…”

“…Space‐flight experiments have indicated that protoplasts have a reduced production of cellulosic compounds and a decreased frequency of cell divisions under the conditions of weightlessness. In addition, protoplast volume has been observed to increase under micro ‐g conditions (Iversen et al 1992; Klimchuk et al 1992; Rasmussen et al 1992, 1994a,b; Iversen et al 1999).…”

Plant cell gravisensitivity and adaptation to microgravity

“…Gray , Helianthus annuus L. , Lactuca sativa L. (all Asteraceae, eudicots; Merkys & Laurinavichius ; Brown & Chapman ; Halstead & Dutcher ; Rasmussen et al . ; Perbal & Driss‐Ecole ; Tripathy et al . ; Kordyum ; Kiss et al .…”

Gravity Research on Plants: Use of Single-Cell Experimental Models

“…It is in particular the ability to regenerate mature plants from protoplast cultures that makes this experimental system interesting for applications in space exploration. Under micro-gravity conditions, cell wall formation in protoplasts isolated from Brassica napus , Daucus carota , and Solanum tuberosum is significantly delayed compared with the control samples at 1- g (Nedukha et al, 1994; Rasmussen et al, 1994, 1992). In particular, the content of structural components such as cellulose and hemicellulose is reduced whereas pectin is unaltered (Nedukha, 1998; Skagen and Iversen, 2000).…”

“…In these studies, peroxidase activity was measured in the regenerating protoplasts and revealed a decrease in enzyme activity compared to the ground control (Rasmussen et al, 1992). Since peroxidase activity is involved in cell wall metabolism and cross-linking of microfibrils, this reduced activity was proposed to provide a possible explanation for the observed slow-down in cell wall regeneration.…”