Effect Of Phosphate Stress On Glycolipid Metabolism And The Function Of Key Gene In Synechococcus

Glycolipids are lipid components that are covalently bonded to monosaccharides or polysaccharides.Monogalactosyldiacylglycerol(MGDG),digalactosyldiacylglycerol(DGDG) and sulfoquinovosyldiaclglycerol(SQDG)are the aboundant natural glycolipids.They are widely found in plants and microalgae.Glycolipids not only serve as structural lipids,which determine the fluidity and integrity of membranes,but also have biological activities in food,pharmaceutical,fuel and cosmetics inducstry.Glycolipids are sensitive to changes in the environment.Reduced phosphate concentration induces glycolipid accumulation and lipid remodeling in cyanobacteria.However,the metabolic mechanism of glycolipids and the physiological significance of glycolipids in resistance to low phosphorus stress have not been studied.Here,the effect of phosphate starvation on glycolipid metabolism and the function of key gene in Synechococcus sp.PCC 7942 were studied.In Synechococcus sp.PCC 7942,the content of glycolipids increased with reduced phosphate concentrations.The glycolipids content increased to as high as 6.13%(of dry cell weight,DCW)and phospholipids(mainly phosphatidylglycerol)decreased to 0.34%(of DCW)after 16 days of cultivation without phosphate.Glycolipids accumulation were mainly attributed to the significant increase of DGDG by 50%and SQDG by 90%.The relative expression levels of glycolipid genes were increased significantly under phosphate starve conditions.DGDG and SQDG could act as complementary lipids for phosphatidylglycerol(PG)to maintain the intergrity of the cyanobacterial thylakoid membrane and improve the photosynthetic activities.Also,the content of C12 and C14 in phosphate-depleted cells was only 28%of the control group(0.04 g L^-1).A notable increase in content(by 48%)of C18:1 was observed in all glycolipids.These changes may contribute to membrane fluidity and photosynthetic activity for basic cell metabolism and phosphate stress adaptation.Lipidomic analyses showed the reduction of PG 18:1/16:0(by 52%)with the increase of DGDG 18:1/16:0(133%)and SQDG 18:1/16:0(245%),strongly suggesting a direct conversion of PG to DGDG and SQDG via intermediate diacylaglycerol.Moreover,the decreasing amount of MGDG 16:1/16:0(22%)was consistent with the increase of free fatty acids(125%)on day 2of phosphate absence,which suggested that MGDG is more likely to provide a pool of fatty acids for synthesis of glycolipids.These results helped us understand the process underlying this glyclipid accumulation and lipid remodeling mechanism in phosphate-starved cyanobacteria.It was also suggested that this is likely one of an important mechanisms to adaptate to phosphate stress conditions for cyanobacteria.Different to higher plants,cyanobacterial MGDG synthesis involves in two-steps reactions.Monoglucosyldiacylglycerol(MGlcDG)not only serves as a precursor for MGDG synthesis,but also participates in stress acclimation.Two genes(synpcc7942?1083 and synpcc7942?0861)related to MGDG synthesis of Synechococcus sp.PCC 7942 were identified.The result suggested that the candidate gene synpcc7942?1083(mgd A)could catelyze MGlcDG synthesis and synpcc7942?0861(mgd E)e Fncoded enzyme responsible for MGlcDG epimerization.The mgd E-suppressed mutant(AE)accumulated MGlcDG(4.2%).After 18 days phosphate starvation,the OD₇₃₀ value(0.468)and chlorophyll content(0.49?g m L^-1)of AE reached to the highest among the other cyanobacteria strains without MGlcDG accumulation.Moreover,the F_V/F_M value of AE decreased more slowly than that of the other strains under phosphate-starved conditions.This result suggested that MGlcDG was involved in phosphate stress adaptation for Synechococcus sp.PCC 7942.A notable increase in contents of 18:1 of MGDG(127%),DGDG(68%),and SQDG(105%)in AE were found during phosphate starvation.However,the expression of?9desaturase(des C)was not higher in AE than that in wild-type during phosphate-starved period.These results suggested that MGlcDG might be involved in the process of fatty acid(FA)desaturation,which contributed to membrane fluidity and cell basic metabolism for stress acclimation in cyanobacteria.The result of complementary experiments of E.coli further suggested that the accumulation of MGlcDG could prompt FA desaturation in E.coli.Therefore,we propose that an overproduction of MGlcDG is responsible for FA desaturation and participates in phosphate stress adaptation in cyanobacteria.In summary,this doctoral dissertation revealed the mechanism of glycolipid accumulation and membrane lipid remodeling under phosphate-starved conditions,identified the function of genes involved in MGDG synthesis in Synechococcus sp.PCC 7942,speculated the important function of MGlcDG in phosphate stress adaptation and developed a high-throughput lipidomic analysis method in Synechococcus sp.PCC 7942.This work will provide a reference for further understanding the metabolic mechanism of cyanobacteria under phosphate stress conditions,and ultimately provide scientific support for efficient biological synthesis of glycolipids.