| Cyanobacteria is the most abundant group of photosynthetic autotrophic prokaryotes on Earth.Synechococcus is a major representative of cyanobacteria and makes important contributions to oceanic carbon fixation,while also serving as an important chassis strain in synthetic biology.Synechococcus carries out photosynthesis through several pigment-protein complexes(PSI,PSII,cytochromeb6f complex,ATP synthase and RuBisCO),converting dissolved inorganic carbon(DIC)into biomass from the ocean.Through active excretion,passive leakage or cell lysis,Synechococcus-derived dissolved organic matter(SOM)is released by Synechococcus and forms an important basis for the biological carbon pump.Both improving the carbon fixation efficiency of Synechococcus and enhancing its application value in synthetic biology involve differential gene expression.Selective RNA processing and stabilization(SRPS)is a post-transcriptional regulatory mechanism in prokaryotes that mainly acts through SRPS-enzymes to resolve conflicts between manipulating genes within operons simultaneously transcribed with differentially expressed genes.Previously,our laboratory has analyzed the molecular mechanisms of SRPS molecules in Escherichia coli.Meanwhile,analysis of published dRNA-seq data from pure cultures of marine microorganisms revealed that multiple operons are potentially regulated by SRPS.For example,it was found that SRPS participates in regulating 32 protein complexes including photosystems Ⅰ&Ⅱ,phycobiliproteins and photoinhibition proteins etc.,based on Prochlorococcus marinus strMIT9313 as an example.Therefore,this study focused on Synechococcus sp.PCC7002(hereinafter referred to as PCC7002),knocked out its SRPS-enzymes using a series of pure culture experiments to explore how SRPS affects the growth rate,photosynthetic pigment content,chlorophyll fluorescence,carbon and nitrogen content and expression of key protein subunits involved in photosynthesis in PCC7002.The main conclusions are as follows:1.Knocking out SRPS-enzymes mainly affects the growth rate of Synechococcus by reducing its iteration speed or changing its growth time.In terms of maintaining the growth time of PCC7002,knocking out endonucleases mostly reduces the photosynthetic pigment content by shortening the growth time,while mutations in exonucleases genes mainly promote their effect on photosynthetic pigment content by prolonging their growth time.Under changes in saturated light intensity,knocking out SRPS mainly shows inhibitory effects which is consistent with changes observed under strong light conditions(PAR≥100 μmol m-2 s-1)after knocking out SRPS-enzymes on photosynthetic oxygen evolution rate and relative electron transfer rate.2.Analysis of SR,HIX and BIX revealed that mutations in exonucleases(RNaseⅡand PNPase)led to a significant increase in SR;all exonucleases(including RNaseJ)knockouts significantly increased SOM bioactivity;three homologous enzymes variations for endonuclease RNaseⅢ showed consistent trends with significantly reduced SOM bioactivity opposite to that seen for exonuclease PNPase:HIX was consistently higher than other algal strains for PCC7002△Ⅲ-2.3.Knocking out SRPS-enzymes resulted in an increase in subunit expression levels for most cytochromeb6f complexes but a decrease in subunit expression levels for most PSⅠ,PSⅡ,RuBisCO and COX.We speculate that all participate in the regulation of photosystems Ⅰ&Ⅱ,cytochrome b6f complex.ATP synthase and RuBisCO.This paper focused on PCC7002 as the research object to explore the impact of SRPSenzymes on photosynthetic carbon fixation efficiency by measuring a series of photosynthetic carbon fixation indicators and conducting qRT-PCR experiments targeting key protein subunits involved in photosynthesis.The simultaneous 5 ’ and 3’ end sequencing process was optimized to lay a foundation for analyzing the molecular mechanisms underlying how SRPSenzymes affects photosynthetic carbon fixation efficiency in PCC7002. |
