Screening And Identification Of Cyanobacteriochromes Interacting Proteins In Nostoc Flagelliforme

Cyanobacteriochromes(CBCRs)are an important class of functional pigment protein photosensitizing light in cyanobacteria and are capable of responding to light signals in the near-ultraviolet to far-red range,and severse light conversion effect similar to plant phytochromes.With the development of molecular biology and genetic engineering technology,the molecular structure,photochemical properties and biosynthesis of CBCRs have made great progress,but little research has been done on its biological functions and light signal transduction mechanism.In this study,on the basis of the complete screening of cyanobacteriochromes,we screening and studying the CBCRs interaction proteins in Nostoc flagelliforme,to provide the basis support of light-signal transduction mechanism and physiological metabolic pathway involved in cyanobacteriochromes.Based on the most photoreceptors are respond to red light in N.Flagelliforme.Cyanobacteriochromes(COO91₀5367,COO91₀5935,COO91₀7954,COO91₀8839,COO91₀0652,COO91₀3972)that respond to red light were selected for screening and identification of interaction proteins.By yeast two-hybrid assay,we screened cDNA library and transcription factor library to obtain the protein interacting with the cyanobacteriochromes.Through the screening,identification,point-to-point validation,and in vitro pull-down experiments,some proteins interacting with CBCRs were preliminary obtained.By screening the cDNA library of N.flagelliforme,the cyanobacteriochromes COO91₀3972,COO91₀7954,and COO91₀5935 obtained some interacting proteins.Some candidate interacting proteins were selected to verify by point-to-point through yeast two-hybrid assay.Result display:COO91₀3972 can interact with multidrug efflux pump subunit AcrA(COO91₀0532),DNA repair protein(COO91₀6633),zinc protease(COO91₀2658)and porin(COO91₀7068).COO91₀7954 was found to interacted with four proteins,which are TPR family proteins(COO91₀0200),FtsH proteins(COO91₀3537),proteolytic enzymes(COO91₀5428),and ATP-dependent proteases(COO91₀3305).Two proteins were screened interacting with COO91₀5935,namely the protein emrA in multidrug efflux system(COO91₀5192)and the Anion permease ArsB(COO91₁1272).By analyzing the CBCRs domain in N.flagelliforme,most of the CBCRs contained a kinase domain,indicating that CBCRs may interact with transcription factors and participate in light signal regulation pathway.Therefore,in this study,six cyanobacteriochromes wered screened of the transcription factor library and finally six transcription factors were found which can interact with the CBCRs.Among them,COO91₀5367 can interact with transcription factors COO91₀3874 and COO91₀0313.COO91₀7954 can interact with transcription factors COO91₀3874,COO91₀0313 and COO91₀7955.Both COO91₀8839 and COO91₀5935 can interact with the transcription factor COO91₀3 874.COO91₀0652 can interact with transcription factors such as COO91₀3874,COO91₀0313,COO91₁0495,COO91₀3149 and COO91₀6808.COO91₀3972 can interact with transcription factors COO91₀3874 and COO91₀6808.COO91₀7954 is located in the upstream adjacent position of its interacting transcription factor COO91₀7955 on the genome,suggesting that they can form two-component signal transduction component.In addition,the transcription factor COO91—03874 can interact with multiple cyanobacteriochromes,and its homologous protein Ar13768(OrrA)in Nostoc sp.PCC 7120 can regulate polysaccharide synthesis,responsing to drought,and osmotic stress.The transcription factor COO91₀0313 belongs to a family of proteins that have functions such as carbon metabolism reaction and stress response.These selected interaction proteins are compatible with the physiological characteristics of N.flagelliforme,such as long-term adaptation to drought,high-ultraviolet radiation and rapid recovery after water supply.Which provides a basis for exploring the light signal transduction mechanism of N.flagelliforme in adapting to extreme environments.It also provides a theoretical basis for exploring the physiological functions and photobiological effects of cyanobacteriochromes.