条斑紫菜早期发育阶段响应盐度胁迫的生理生化过程分析

The early development of organisms is usually accompanied with the adjustment of metabolism. This regulation makes the cell to be better acclimatizing to the environment which it will experience in future. Provision of ATP and NAD(P)H is believed to be the most important reaction of development. Pyropia yezoensis (Py. yezoensis) belongs to an important economic seaweed. The thallus of Pyropia shows strong resistance but conchocelis is susceptible to stress conditions. As the transition period of these two generations, development of conchospore is believed to be the key stage of germination of stress resistance. The project focuses on early development stages of Py. yezoensis to explain the relationship between cell wall formation and the emergence of stress resistance. Thus, the photosynthesis parameters of different development stage conchospores under different salinity stress will be determined and the effect of salinity on the livability be assessed. The occurrence of the cyclic electron flow (CEF) with development of Py. yezoensis conchospores is determined by the aids of inhibitors. During the process, the potential involvement of the oxidative pentose phosphate pathway (OPPP) will be analyzed through determination of enzymes activities. Based on the transcriptome analysis of the specified samples, the molecular mechanism of stress tolerant ability related to the metabolism regulation is expected to be illuminated. The completion of this project will not only contributes to intensively understanding of the OPPP mechanism on stress tolerance and regulation of metabolism, but also provides theoretical guidance for the cultivation of macroalge in future aquaculture.

生物体早期发育往往伴随着代谢的调整，使得更有利于后续生存生长。能量与还原力供给是这个过程最为重要的生化过程。条斑紫菜是重要的经济海藻，叶状体具有极强抗逆性，丝状体则基本不表现对逆境的适应，作为连接这两个阶段的壳孢子附着和细胞壁的形成，是抗逆性状获得的关键时期。本项目聚焦壳孢子发育过程中悬浮壳孢子、附着壳孢子以及紫菜幼苗（3~10个细胞）不同发育阶段，研究细胞壁形成与抗逆机制获得的关系。分别研究不同盐度条件对光合作用参数及后续生长发育影响；在电子传递抑制剂条件下，研究环式电子传递随发育阶段不同而改变的规律；通过测定各阶段磷酸戊糖途径（OPPP）关键酶的活性，评估该途径与环式电子传递相关性；通过转录组测定与分析，解释基于代谢调控的OPPP途径对条斑紫菜抗逆性状获得的分子机制。项目的完成，有助于丰富人们对OPPP途径在条斑紫菜抗逆性状及代谢调控方面的认识，也为抗逆品系的培育提供了新的思路。

条斑紫菜是重要的经济海藻，叶状体具有极强抗逆性，丝状体则基本不表现对逆境的适应，作为连接这两个阶段的壳孢子附着阶段，是抗逆性状获得的关键时期。本项目通过实验室内培养条斑紫菜贝壳丝状体，摸索不同环境因子（无机碳、光质）对丝状体形成壳孢子囊枝的影响以及结合壳孢子放散的条件，获得了条斑紫菜悬浮壳孢子，附着壳孢子以及3-10细胞紫菜幼苗，利用荧光白染色的方法观察了壳孢子细胞壁形成过程。测定了其早期不同发育阶段在盐度胁迫条件下Fv/Fm、ETRI、ETRII等光合参数的变化，进一步通过光合参数变化结果重点关注了附着壳孢子在不同盐度胁迫条件下关键代谢途径中的关键酶活（G6PDH、6PGDH、cGAPDH）、还原力(NADPH、NADH)以及光合电子传递之间的关系。研究结果表明悬浮壳孢子在盐度为4%时几乎没有光合活性，而附着壳孢子在盐度为18%时仍具有一定的光合活性，但光合活性值小于3-10细胞小紫菜，小紫菜叶状体已具备较强的抗逆能力。附着壳孢子光系统II的活性在18%盐度时，ETRII的值几乎为零，而光系统I仍具有一定的活性，在添加DCMU抑制剂时测得附着壳孢子光系统I活性随着盐度的升高而上升，进一步通过添加DBMIB抑制剂发现光系统I的活性值很低并没有一定的趋势，结果表明盐度胁迫条件下附着壳孢子细胞内光合活性来源于光系统I的环式电子传递而不是光系统II。在盐度胁迫条件下（4%-12%）OPPP关键酶G6PDH、6PGDH酶活及还原力NADPH呈上升的趋势，而糖酵解途径关键酶活cGAPDH以及还原力NADH随盐度的升高呈下降趋势。在18%盐度时无论是OPPP还是糖酵解途径中的酶活和还原力均呈较低的值，说明附着壳孢子此阶段活性较差。研究阐明条斑紫菜抗逆的特性确实是发生在早期发育阶段，并且和壳孢子附着密切关联，OPPP途径产生的还原力为附着壳孢子环式电子传递提供电子来源。本项目的完成不仅系统的研究了紫菜光合活性和糖代谢途径的协同抗逆机制，而且合理地解释了壳孢子不同发育阶段物质和能量的供给，同时为生产过程中紫菜抗逆特性的应用提供理论指导。