Galactinol synthase gene 5 (MdGolS5) enhances the cold resistance of apples by promoting raffinose family oligosaccharide accumulation

Low-temperature stress is a limiting factor affecting the safe overwintering and stable production of apples. Galactinol, produced by galactinol synthase (GolS), is an important plant cryoprotectant. This study showed for the first time that exogenous spraying of apple saplings with 100 mg mL^-1 galactinol could effectively alleviate the damage from low-temperature stress. Further, we found that transgenic apple callus and tobacco overexpressing MdGolS5 showed strong cold tolerance. Specifically, the activities of antioxidant enzymes such as superoxide dismutase and GolS in transgenic tobacco overexpressing MdGolS5 increased under low-temperature treatment at -2 °C, and the contents of malondialdehyde, superoxide anion, and hydrogen peroxide were significantly lower than those of wild type tobacco. Moreover, large amounts of proline, galactinol, and raffinose were accumulated. In addition, the expression levels of cold-responsive genes MdCBF1, MdCBF2, MdCBF3, and MdCOR47 were significantly up-regulated in transgenic tobacco, further confirming the important role of MdGolS5 in regulating plant cold adaptation. In summary, this study not only revealed the direct effect of exogenous galactinol on the low-temperature protection of apple saplings for the first time, but also explored a new mechanism of raffinose family oligosaccharides anabolism in plant low-temperature adaptation through overexpression of MdGolS5. These results provide a theoretical basis for the genetic improvement of apple cold resistance.