DataSheet_1_Heterologous overexpression of the cyanobacterial alcohol dehydrogenase sysr1 confers cold tolerance to the oleaginous alga Nannochloropsis salina.docx

Temperature is an important regulator of growth in algae and other photosynthetic organisms. Temperatures above or below the optimal growth temperature could cause oxidative stress to algae through accumulation of oxidizing compounds such as reactive oxygen species (ROS). Thus, algal temperature stress tolerance could be attained by enhancing oxidative stress resistance. In plants, alcohol dehydrogenase (ADH) has been implicated in cold stress tolerance, eliciting a signal for the synthesis of antioxidant enzymes that counteract oxidative damage associated with several abiotic stresses. Little is known whether temperature stress could be alleviated by ADH in algae. Here, we generated transgenic lines of the unicellular oleaginous alga Nannochloropsis salina that heterologously expressed sysr1, which encodes ADH in the cyanobacterium Synechocystis sp. PCC 6906. To drive sysr1 expression, the heat shock protein 70 (HSP70) promoter isolated from N. salina was used, as its transcript levels were significantly increased under either cold or heat stress growth conditions. When subjected to cold stress, transgenic N. salina cells were more cold-tolerant than wild-type cells, showing less ROS production but increased activity of antioxidant enzymes such as superoxide dismutase, ascorbate peroxidase, and catalase. Thus, we suggest that reinforcement of alcohol metabolism could be a target for genetic manipulation to endow algae with cold temperature stress tolerance.

温度是藻类及其他光合生物生长的重要调控因子。当生长温度偏离最适区间时，会通过活性氧（reactive oxygen species, ROS）等氧化合物的积累对藻类造成氧化胁迫。因此，可通过提升藻类的氧化胁迫抗性，实现其温度胁迫耐受性的强化。在植物中，乙醇脱氢酶（alcohol dehydrogenase, ADH）已被证实与低温胁迫耐受性相关，可诱导抗氧化酶的合成以抵御多种非生物胁迫引发的氧化损伤。但藻类中ADH能否缓解温度胁迫，目前仍鲜有研究。本研究以单细胞产油藻类盐生微拟球藻（Nannochloropsis salina）为材料，构建了异源表达sysr1的转基因株系；sysr1编码集胞藻（Synechocystis sp. PCC 6906）中的ADH。为驱动sysr1的表达，本研究使用了从盐生微拟球藻中克隆得到的热休克蛋白70（heat shock protein 70, HSP70）启动子，该启动子的转录水平在低温或高温胁迫培养条件下均显著上调。经低温胁迫处理后，转基因盐生微拟球藻细胞的低温耐受性优于野生型细胞，其ROS生成量更低，且超氧化物歧化酶（superoxide dismutase, SOD）、抗坏血酸过氧化物酶（ascorbate peroxidase, APX）及过氧化氢酶（catalase, CAT）等抗氧化酶的活性更高。综上，本研究提出，强化乙醇代谢可作为遗传改造靶点，使藻类获得低温胁迫耐受性。