| Coccolithophores are considered to be one of the most productive calcified organisms on earth and it accounts for one-third of the annual ocean calcium carbonate production of 5 billion tons.The scale of its photosynthetic carbon sequestration(biological carbon pump)and the characteristics of releasing carbon by calcification(carbonate reverse pump)make it play a pivotal role in regulating marine biogeochemical cycles and global climate.A large amount of man-made CO2 emitted through the use of fossil has caused global climate change and ocean acidification.At the same time,continuous ocean acidification may aggravate the iron limitation to phytoplankton.Increasing ocean acidification and iron limitation have respectively caused serious threats to the calcification and growth of coccolithophores,what may affect the feedback adjustment of coccolithophores and global climate,and are worth studying.However,it is not clear what effect these two factors will have on coccolithophores.There are many contradictory results in the effects of p H/p CO2 on the calcification of coccolithophores,and the internal mechanism lacks in-depth analysis.Little is known about the effects of iron limitation on their photosynthetic physiology and the possible role of calcification in it.Emilinia huxleyi is the most abundant coccolithophores in the world and is easily cultured.It is the most intensively studied member of the coccolithophores.In this paper,PERU41 and PERU46,which were isolated from the Peru upwelling,with different genotypes and different calcification phenotypes,were used as the research object to explore the differential response and possible adaptation mechanism under low p H and iron limitation conditions.Relative to the average p H of the world ocean(8.1),the seawater in this area is slightly acidic(p H=7.9,the lowest is 7.65),although it benefits from the rich nutrients brought by upwelling,but periodic El Ni?o can seriously reduce the availability of iron in Peruvian upwelling waters.Therefore,these two strains of E.huxleyi are ideal models for studying the effects of low p H and iron limitation in this paper.Two strains of E.huxleyi were cultured under a set p H gradient(8.2,7.8,7.5,and7.2)until the cell density reached 5 × 104 cells/m L,and then the differences in parameters of calcification and gene expression of transcription levels were measured and compared.The study found that there were significant differences in calcification level and growth rate between two different genotypes.The results of scanning electron microscope,carbonate chemistry and particulate carbon all support that PERU41 with higher calcification levels.The possible calcification-related genes in PERU41 are upregulated and there are fewer differentially expressed genes between different p Hs,indicating that it maintains calcification by regulating gene expression and is more tolerant to low p H.The higher growth rate of PERU46 and the up-regulation of DNA replication-related genes proved its advantage in proliferation.This paper speculates that the two genotypes may have different survival strategies during long-term adaptation,especially under low p H stress: the PERU41 of ? clade may belong to K strategist that with higher calcification level and lower growth rate,and more energy is used for calcification;the PERU46 of ? clade belongs to r strategist that with lower calcification level and higher growth rate,and tends to use more energy for proliferation.Among them,there may be an energy trade-off mechanism in PERU41.Low p H may increase the demand for energy,while maintaining calcification requires more energy,so an increase in energy metabolism occurred.At the cost of sacrificing part of the reproductive capacity,a large number of genes involved in energy-coupled transmembrane transport are up-regulated.However,PERU46 still uses energy for proliferation under low p H conditions and up-regulates "DNA repair" related genes to overcome the negative effects of low p H on proliferation.Finally,in view of the influence of genotype and p H on the calcification phenotype,this thesis compares the differentially expressed genes and their functions,and combined with previous studies,to propose a cellular model of the calcification process.At the same time,possible calcification-related genes are listed.ABC transporter,Vesicle H+-ATPase,Stx1-4,VAMP7 and SNARE homologues are worth noting in the follow-up study.The five iron concentrations(0,50,100,500,1000 n M)had no significant effect on the growth of the two strains,but the specific growth rate of PERU41 was significantly higher than that of PERU46 at 0 n M.When iron was limited,the cell size and volume decreased,and the maximum electron transfer rate and apparent photosynthetic efficiency decreased.And the maximum value appeared at 100 n M.Periodic El Ni?o can cause dramatic changes in the bioavailable iron in the Peru upwelling,which makes E.huxleyi achieve long-term adaptation under the conditions of high and low iron.However,the results of pigment and chlorophyll fluorescence showed that there were differences in their photoprotective mechanisms under iron restriction.PERU41 has a significantly higher carotenoid content than PERU46 at low iron,while non-photochemical fluorescence quenching(NPQ)is the opposite.It is speculated that PERU41 achieves photoprotection through the lutein cycle and highly calcified coccoliths,while PERU46 uses more other forms of non-photochemical fluorescence quenching.The possible response of the calcification of E.huxleyi to iron restriction was further studied by coupling iron(high iron 100 n M,low iron 0 n M)with light(high light 300 ?mol/m2/s1,low light 20 ?mol/m2/s1).The effect of light on photosynthesis of coccolithophores was found to be greater than that of iron.Under low light(20 ?mol/m2/s1),cells may increase light capture by increasing photosynthetic pigments,while non-photochemical fluorescence quenching is significantly increased under low-light and iron-deficiency coupling conditions.Compared with low light conditions,pigments are significantly reduced under high light.PERU46 maintained higher maximum photochemical efficiency of PSII and effective quantum yield of PSII,the maximum electron transfer rate and apparent photosynthetic efficiency under high-light and iron deficiency conditions,indicating its tolerance to high light,and the existence of a possible relatively low iron content in the photosynthetic structure or a compensatory mechanism for maintaining electron transport in cells.However,The PIC/POC of PERU41 increased significantly under high-light and iron deficiency conditions,and Fv/Fm and POC content decreased significantly under high light,indicating that photosynthesis was inhibited under high light condition.The two strains had different tolerance to light.The environmental conditions combining iron limitation with high light did not stimulate the enhancement of calcification to realize the photoprotection of cells as speculated.On the contrary,the content and productivity of particulate organic carbon of PERU41 and PERU46 decreased in response to iron limitation under low light and high light,respectively.Whether the calcification in calcification is directly related to iron needs to be further explored.Through the above research,it is found that different genotypes of E.huxleyi have different adaptation strategies to cope with ocean acidification and iron limitation,suggesting the strong and diverse potential adaptability of E.huxleyi and even more coccolithophore groups to climate change.This study provides new molecular evidence for elucidating the calcification mechanism and iron-deficiency adaptation mechanism of coccolithophores,and provides new ideas for how coccolithophores participates in and feedback-regulates the biological carbon cycle,which has certain scientific significance. |
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