The Effects Of Nitrogen Limitation On Carbohydrate Accumulation In Cyanobacteria And Its Regulation Mechanisms

The cyanobacterial blooms caused by eutrophication is a global environmental problem,and their occurrence and disappearance mechanisms has become a hot topic.Nutrient limitation is one of the important factors affecting the proliferation of cyanobacteria.Our previous studies have found that nitrogen limitation could inhibit the growth of Microcystis and induce intracellular carbohydrate accumulation,furtherly increase the specific density and promote the sinking of Microcystis.The sinking has a negligible contribution to the disappearance of cyanobacterial blooms.However,at present,there are few studies on the mechanisms of carbohydrate accumulation in Microcystis under nitrogen-limited conditions.In this study,the physiological responses of Microcystis to nitrogen limitation were investigated,including the changes of cell components and the response of photosynthetic system,and the multi-omics method was used to explore the cell regulation mechanism for the carbohydrate accumulation.This study will help to provide theoretical support for clarifying the disappearance mechanisms of Microcystis blooms.The main conclusions of this study are as follows:(1)Initial nitrogen sources could affect the carbohydrate accumulation in Microcystis cells after the depletion of nitrogen source.Compared to initial addition of ammonia and urea,initial addition of nitrate significantly promoted the carbohydrate content of Microcystis,which was rapidly increased to 23.81% on Day 6 and to 68.90%on Day 9.(2)Under nitrogen-limited condition,the values of Fv/Fm,? and ETRmax of Microcystis were significantly increased,suggesting that the activity of photosynthetic system II(PSII)was enhanced,the original reaction of photosynthesis was promoted,and the photosynthetic electron transfer process was strengthened.At the same time,the content of pigment in Microcystis decreased slightly and the percentage of pigment was different from that in nitrogen-sufficient Microcystis.The percentage of chlorophyll and carotenoids decreased with time.The percentage of phycocyanin increased to 18.10% of the maximum,then decreased to the lowest value of 14.30%.The percentage of allophycocyanin also reached the maximum at 12 h,which was30.64%.The percentage of phycoerythrin was 47.43% at 12 h.Under nitrogenlimitation condition,the net photosynthetic oxygen evolution of nitrogen-limited Microcystis decreased significantly in Microcystis;the value of nitrogen-sufficient Microcystis changed little,and the difference between the lowest value and the highest value was within 23%.The change of respiration rate showed the opposite trend,and the nitrogen-sufficient Microcystis decreased significantly,and the respiration rate of the nitrogen-limited Microcystis changed little during the experiment.(3)The proteomic results showed that in Microcystis under nitrogen limited condition for 12 h,there were 441 proteins upregulated and 312 proteins downregulated,while,in Microcystis under nitrogen limited condition for 48 h,there were 504 proteins upregulated and 451 proteins downregulated.KEGG analysis showed that in the photosynthesis pathway,the proteins involved in the PS II reaction center was upregulated,indicating the ability of Microcystis to capture light energy after nitrogenlimitation treatment.The activity of PS II excretion enhancer protein 1 and PS II excretion enhancer protein 2 was down-regulated,which further led to the decrease of oxygen release rate of Microcystis cells.The results were consistent with the results of the above physiological responses.The proteins involved in the carbon fixation pathway were down-regulated,indicating that the synthesis of carbohydrate did not increase.The proteins involved in the glycolysis pathway were significantly downregulated.Combining with the result of down-regulation of the pathway related to pyruvate conversion to protein and fat,this indicates that the accumulation of carbohydrates in Microcystis is closely related to the consumption of carbohydrate under nitrogen limitation.What's more,it is preliminarily considered that the downregulation of glycolysis pathway is an important reason to induce carbohydrate accumulation in algae.