Accumulation of Glycinebetaine in Chloroplasts Provides Osmotic Adjustment During Salt Stress

Abstract

Glycinebetaine was determined in leaves and in isolated chloroplasts of spinach (Spinacia oleracea) by nuclear magnetic resonance spectroscopy. Some leakage of glycinebetaine from the chloroplasts occurred during the isolation so the concentration in chloroplasts in vivo could be up to 1.5 times higher than that measured in isolated chloroplasts. It was demonstrated that any contamination of the chloroplast preparations by glycinebetaine originating from other cellular compartments or from broken chloroplasts would have amounted to less than 10% of the measured values. Leaf osmotic potential of salt-stressed plants was -2.09 MPa compared to -0.91 MPa in non-stressed controls. This was accompanied by a sixfold increase in glycinebetaine content in the leaf but the levels of choline and proline were not increased. In chloroplasts isolated from control leaves the calculated glycinebetaine concentration was 26 mM which was 10-fold higher than the concentration in the leaf as a whole but only contributed 7% of the osmotic potential of the chloroplast. Chloroplasts from salt-stressed plants contained up to 300 mM glycinebetaine which was 20 times the concentration in the leaf as a whole. The glycinebetaine concentration in chloroplasts from salt-stressed leaves was equivalent to an osmotic potential of -0.75 MPa and this contributed 36% of the osmotic potential of the chloroplast and 64% of the decrease in osmotic potential induced by salt stress. At least 30-40% of the total leaf glycinebetaine was localized in the chloroplast. The results demonstrate that glycinebetaine accumulates in chloroplasts to provide osmotic adjustment during salt stress and provide support for the hypothesis that glycinebetaine is a compatible cytoplasmic solute which may be preferentially located in the cytoplasm of cells.