Association of the cold-hardiness of Chorispora bungeana with the distribution and accumulation of calcium in the cells and tissues

Chorispora bungeana Fisch. & C.A. Mey (Chorispora bungeana) is a rare alpine subnival plant species that is highly capable of resisting freezing environment. In this paper, we attempted to explore a possible mechanism of its adaptation to the alpine subnival environment. The results of Ca2+ level measured by energy-dispersive X-ray detector (EDS) showed that the Ca2+ content in the wild species was higher than that of regenerated plants, and chilling induction at 0 °C could increase Ca2+ contents in the regenerated plants. Further, the distribution and accumulation of Ca2+ in tissues was investigated by X-ray micro-domains analysis with scanning electron microscope (SEM), the results showed that Ca2+ levels were different in various tissues and organs of the wild species. However, Ca2+ levels did not show significant difference among tested tissues of the regenerated plants. Laser-scanning confocal microscopy (LSCM) revealed that, after 10th day of the chilling induction at 0 °C, the free Ca2+ levels increased in the mesophyll protoplasts from the regenerated plants. In addition, the results of electric conductivity (EC) suggested that the chilling induction initially affected the permeability of cellular membrane, and it was restored to stable level with extending treatment. The mitochondria of the cells also maintained its normal function after the chilling induction by Rhodamine-123 detection. Taken together, the membrane remained its integrity and its mitochondria was functional at the condition of the chilling induction, which was essentially the requirement for alive cells as well as its whole entire plants. These findings suggested that distribution and accumulation of Ca2+ connected with the cold-hardiness of C. bungeana were different among the various tissues and organs in the wild species. Also, these results demonstrated that the difference in Ca2+ distribution was a response of C. bungeana in adapting itself to the alpine subnival environment and the accumulation of Ca2+ could play an important role in its active cold-hardiness.