Effects Of Chilling Stress On Protein And Related Gene Expression In Chloroplasts Of Sugarcane

Sugarcane is the main sugar crop in China and the world, Guangxi is the main sugarcane producer in China, and cane sugar industry is one of the important economic pillars in Guangxi. Sugarcane is a thermophilic crop, but in recent years, extreme low temperature weather occurred frequently, which seriously affected the sugarcane production, and resulted in huge economic losses.The present study employed the strong cold resistant sugarcane variety GT28and weak cold resistant variety ROC22as the plant materials. Two treatments, chilling stress (0-4℃) and no stress control (28℃) were designed. Changes of chloroplast protein expressions were analyzed using one-dimensional and two-dimensional electrophoreses at different time of treatment. Differential expressions of the chilling stress related genes were analyzed using real-time PCR technique. The effects of chilling stress on sugarcane chloroplasts at protein and mRNA levels were discussed to provide a reference for further investigation on molecular mechanism of chilling stress in sugarcane. The main results were as follows.1. The results of one-dimensional electrophoresis showed that, after chilling stress, the chloroplast protein showed a significant change. For two sugarcane varieties, the relative content of chloroplast protein was decreasing with the chilling time extension. The protein bands of39.92kDa,32.95kDa and22.87kDa were down-regulated obviously. The results of two-dimensional electrophoresis showed that, for two sugarcane varieties, the down-regulated proteins were significantly more than the up-regulated ones after chilling stress, and most of the protein points showed obvious degradation at24th day of chilling stress treatment. Ten obvious changed protein spots were selected for mass spectrometry analysis, and the results showed that these changed protein spots are all involved in photosynthesis including light energy absorption, electron transfer and energy synthesis. This indicated that low temperature stress hampered the photosynthesis in chloroplasts, which would further inhibit the photosynthetic efficacy, and then hinder the growth of young sugarcane plants.2. Three different protein gene fragments were successfully cloned, and they are ATP synthase subunit gamma gene fragment atpC, chloroplast23kDa polypeptide of photosystem Ⅱ gene fragment PsbO and oxygen-evolving enhancer protein1gene fragment PsbP. The sequencing results showed that the fragment length was937bp,996bp and721bp, respectively. The fragment of oxygen protein1is its complete open reading frame, which has been registered in the GenBank database with the accession number JQ898540.3. Expressions of the genes were analyzed using real-time PCR, and the results showed that, chilling stress suppressed the expression of atpC gene, but induced the expressions of PsbO and PsbP genes. And two different sugarcane varieties express differently, and the expression of genes had not too much correlation with sugarcane cold hardiness. And these results suggested that the changes are different at the mRNA level from the protein level, reflecting the complexity of the plant response to chilling stress in sugarcane.