Functional Mechanism Analysis Of Ips1,A Protein Stabilizing Photosystem ?,Under High Temperature Stress In Cyanobacteria

The photosystem?PS?I is located in thylakoid membrane and is involved in linear electron transfer,cyclic electron transport and water-water cycle.PSI plays an important role in accumulation of organic compounds and elimination of greenhouse gases.In general,absence of PSI results in complete loss of photosynthesis.Therefore,various mechanisms for protecting PSI have been formed during evolution.One of the important protection mechanisms for cyanobacterial cells under stress conditions is the acclimation that maintains the stability of the PSI.In recent years,some proteins and polypeptides are found to be essential to regulate stabilization and degradation of PSI under various stress conditions,and they play important roles in adaptation to stress environments of cyanobacterial cells.However,the understanding of these proteins is still very limited.In particular,relevant proteins haven't yet been identified under high temperature stress.In this thesis,a new component that named as Ips1,which affected the structure and function of PSI under high temperature,has been successfully identified via screening the transposon tagged mutant library of Synechocystis sp.strain PCC 6803.How this protein affected PSI complex under high temperature stress were conducted in this thesis and related results are shown as follows:?1?Through screening the transposon tagged mutant library of Synechocystis sp PCC 6803,we isolated and identified two mutant strains that were unable to grow under high temperature but grew similarly to WT under growth temperature conditions.Through inverse PCR and other tests,we found that two mutant strains were inserted into the same gene ips1.The results of chlorophyll fluorescence analyses indicated that the mutation of ips1 significantly reduced the activity of PSI under high temperature,but under normal growth temperature,the activity of PSI in?ips1 was similar to the WT.Further by adding electron donor and receptor for PSI complex,the activity of PSI in mutant strain was not recovered.Therefore,we obtained a mutant strain with impaired PSI activity under high temperature.?2?Through analyzing the photosynthetic complexes of major changes in the thylakoid membranes using blue native polyacrylamide gel electrophoresis?BN-PAGE?,we found that deletion of Ips1 specificity affected PSI complexes under high temperature,but the deletion did not influence other photosynthetic complexes,including PSII,Cytb₆f,NDH-1 and ATP synthase.Further,it was found that the accumulation of PSI subunit was not induced by high temperature by analyzing the expression of all these photosynthetic complexes.Therefore,we concluded that the absence of Ips1 resulted in instability of the PSI complexes under high temperature,thus damaging the PSI activity.?3?With the aid of bioinformatics analysis,we found that Ips1 contains a CDC37domain that is indicated as a heat shock protein.Afterwards,through the method of immunoblotting,we found that Ips1 was evidently induced under high temperature condition.These results imply that Ips1 interacts with PSI,which is important to maintain the structure of PSI under high temperature condition.Further,the results of sucrose gradient separation and western blot indicated that Ips1 had a co-migration with the PSI complex.In conclusion,we successfully identified a protein,Ips1,and found that the interaction between Ips1 and PSI is crucial for stabilizing the structure of the PSI,keeping the activity of PSI and maintaining the survival of the cyanobacterium Synechocystis 6803 cells under stress conditions.These results will deepen the understanding of the adaptive and protective mechanisms to high-temperature stress in cyanobacterial cells,and provide important theoretical references for understanding the adaptation of higher plants to high temperature stress.