Ecological Distribution Of Proteorhodopsin Gene In Typical Marine Environments

Proteorhodopsin gene was recently discovered in the ocean as a new member of the rhodopsin family.It is a retinal-binding integral membrane protein which can take light to establish a proton gradient and produce ATP for cell consumption. Proteorhodopsin has been found in many marine bacterial groups,mainly fromα-andγ-proteobacteria originally,and also reported in Bacteroidetes,Euryarchaea,β-proteobacteria and Planctomycetales lately.Proteorhodopsin plays an important ecologcial role as a new path for biological utilization of light energy in the ocean, however,its distribution characteristics in different marine environments are still unclear.To investigate the diversity and distribution of proteorhodopsin gene in the global ocean,degenerate primers were designed based on sequences from the environmental genomic library of the Sargasso Sea.More than 300 sequences were retrieved from the Pacific Ocean,the Atlantic Ocean,the Indian Ocean,the Baltic Sea and the China seas.These sequences were mainly from Proteobacteria,others from Bacteroidetes and Archaea according to the phylogenetic analysis,representing a broad diversity. The discrepancy between proteorhodopsin genotypes from different marine regions were probably related to the environmental factors such as nutrients,chlorophyll concentration and salinity,as revealed by the Canonical Correspondence Analysis.Bacteroidetes,an important bacterial group in the marine environment,is usually abundant in the seawater and plays an important role in mineralization of organic matters.So far,five proteorhodopsin sequences have been identified in culturable bacteroidetes,which are divergent from those in other marine groups.Based on these 5 sequences,a primer set specifically targeting proteorhodopsin genes in Bacteroidetes was designed,and a detailed diversity of this gene was investigated in the East and South China seas.Besides,similar proteorhodopsin sequences were also retrieved from the Global Ocean Sampling(GOS) metagenomic database using our sequences as queries,which helped us to understand the distribution of this gene in other marine environments.The results showed a high diversity of proteorhodopsin genes in Bacteroidetes in the China seas,whose similar sequences spreaded around many other marine regions.The phylogenetic analysis indicated that temperature might be an important factor affecting the distribution of different proteorhodopsin genes in Bacteroidetes in the ocean.Sequences from the East China Sea and the North Atlantic coastal water were found similar to each other,which might represent proteorhodopsin genotypes adapting to the temperate seawater;sequences from the South China Sea and the tropical oceans were found similar to each other,which might represent proteorhodopsin genotypes adapting to the warm water; proteorhodopsin from two flavobcterial strains isolated from the Antarctica might represent the genotypes adapted to the cold water.Besides temperature,other enviromental factors such as trophic condition and salinity might also affect the diversity of this gene in the ocean.By grade filtration,both free-living and particle-attached Bacteroidetes were found to carry this gene and different genotypes were found between the two assemblages.To understand the abundance of proteorhodopsin genes in Bacteroidetes in different marine environments,real-time quantitative PCR method was applied to investigate the quantity of this gene in the East and South China seas.Over twofold more abundant proteorhodopsin genes in Bacteroidetes were found in the offshore waters than the nearshore waters close to the estuary,implying that inheritance of this gene could be important for Bacteroidetes living in the oligotrophic environment.On a verticle profile in the northern South China Sea,the abundance of proteorhodopsin genes in Bacteroidetes decreased rapidly with depths,probably related to the gradient of light intensity along the depths.