| In recent years,industrial and agricultural production led to a sharp increase in anthropogenic emissions of ammonia to the whole ecosystem,which has brought a lot of negative effects.Increased ammonia emissions results in the increasion of ammonia concentration in freshwater,which trigger frequent outbreaks of algal blooms.Previous studies showed ammonia could reduce the photosynthetic efficiency in photoautotropic organisms(such as algae).Cyanobacteria ususally are the dominant algae in eutrophication water(contain higher content of ammonia),which can tolerate relatively high concentration of ammonia.To reveal the mechanism of high ammonia tolerance in cyanobacteria,it can provide a theoretical basis for the further understanding of the internal mechanisms of cyanobacterial blooms.Previous transcriptional profiling showed that high light inducible protein family(Hlips)significantly increased upon ammonia treatment,which suggested that this gene family may play an important role in resisting to ammonia toxicity.Hlips is a class of small chlorophyll binding protein family,which is widely distributed in cyanobacteria,involving in the formation of the photosynthetic systems.In order to explore the function of Hlips involved in the ammonia tolerance,we knockout hliA/B/C/D gene respectively.By comparing the growth of wild-type and muants with treatment of various concentrations of ammonia,single knocked-out mutants are more sensitive to ammonia toxicity than the wild-type strain,and hliB-is the most sensitive to ammonia toxicity,which indicated hliB played more important roles than the other three hlips genes in the tolerance of ammonia toxicity.77K results showed that PSI signal(at 720 nm)of single knocked-out mutants are lower than the wild-type strain,and PS? signal(should be 685 nm)of mutant strains were move to 682 nm This result indicated that Hlips invovled in maintaining the stability of PSI and PS? complex during ammonia treatment.Meanwhile,the content of PsbO(OEC small peripheral peptide)protein in the thylakoid membrane of hliA-and hliB-mutants are much less than wild-type and other two single knocked-out mutants after ammonia treatment.It indicated that HliA/B can maintain the stability of OEC small peripheral peptide during ammonia treatment.Through comparing the PS? repair cycle among four triple knocked-out mutants,it showed that only hliB-C-D-and hliA-C-D-mutants have the PS?repair capacity under ammonia treatment,which indicated HliA/B were invovled in the precess of PS? repair cycle.Furthermore,with exposure to 3 mol L-1 NH4Cl for 24 h,hliA-B-double mutant showed no PS? repair cycle,while the PS? repair of hliCD?double mutant still existed,which indicated HliA/B were essential for maintaining the PS? repair.To investigate HliA/B involved either in the D1 protein degradation or synthesis,the linconmycin were added in the wild-type,hliA-B-and hliC-D-cultures,and detected the amount of D1 protein in three strains during different period of ammonia treatment.The result showed that the amount of D1 protein in hliA-B-mutant were kept constant,and it decreased obviously in the hliCD-mutant and wild-type strains,which indicated HliA/B played important-role in the PS? repair cycle through involving in the D1 protein degradation.Moreover,the values of non-photochemical quenching(NPQ)of hliC-D-were obviously lower than hliA-B-during ammonia treatment,which indicated HliC/D played more important roles in the protection against PS? damage through quenching redundant energy excitation on photosystems centre than HliA/B. |
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