Genome Sequencing And Proteomic Research Of The Oleaginous Microalga Chlorella Protothecoides

Chlorella protothecoides can grow in both autotrophic and heterotrophic conditions.In heterotrophic cells, the oil content reached to50%, the cell density was increasedmore than20-fold. The high oil yield of C. protothecoides makes it an importantresource for biodeisel production. In order to eatablish the platform for molecularresearch and explore methods to improve oil production of microalgae, we sequencedthe genome of C. protothecoides and investigated the proteomic changes betweenautotrophic and heterotrophic cells.The genome of C. protothecoides was sequenced by Solexa Paird-End, Solexa MatePair and Roche454method, the coverage of the clean data was120x. After assembly406scaffolds (>100bp) were obtained, and the N50was24. The size of the genome is22.9Mb, and7.039genes are predicted. By comparative analysis of the genome, threemain characteristics were found. Firstly, the C. protothecoides has a small genome insize, and about half of what its close relative Chlorella variabilis NC64A has.Furthermore, it encodes the fewest number of genes among all sequenced green algae,and the decrease of total gene is partly because the decrease of genes with the similarfunction. Secondly, a family of hexose/H+cotransporter that restricts to Chlorella wasfound, and there are three such genes in C. protothecoides, which may participated inthe transform of glucose across the plasma membrane. Thirdly, C. protothecoides is theonly eukaryotic green algae that lack nitrate transporter, but represented a biaseddistribution of amino acid transporters, indicating that C. protothecoides prefer to useamino acid as nitrogen source.The proteins of autotrophic and heterotrophic C. protothecoides were identified byLC/MS/MS.1,931proteins were identified which accounted for27.4%of the totalpredicted proteins. Compared with autotrophic cells,206proteins were up regulatedand288proteins were down regulated. The functional analysis of the differentlyexpressed proteins revealed the changes of the metabolic pathways. The process ofglycolysis, TCA cycle and the synthesis of oil are all enhanced, meanwhile, thephotosynthesis and carbon fixation were inhibited. The results illustrated the transitionof energy biosythesis from photophosphorylation to oxidative phosphorylation in heterotrophic cells. The enzymes in glycolysis are the most abundant proteins inheterotrphic cells. High glycolytic capacity ensured the cell to meet the demand ofprecursors for fatty acid synthesis, and the increase of the fatty acid synthesis enzymesdirectly lead to oil accumulation. The proteins of PP2C and SnRK2in abscisic acid(ABA) signal transduction were also upregulated in heterotrophic cells. ABA may playan important role in heterotrophic growth and oil accumulation.This study established the molecular research platform of C. protothecoides throughthe completement of genomic sequencing and proteomic research. Meantime, the studyrevealed the characteristics of C. protothecoides genome, and the changes of metabolicpathways in autotrophic and heterotrophic cells were illustrated. The results indicatedthe mechanism of oil accumulation in heterotrophic cells. Moreover, the possiblesignal transduction and regulatory proteins were found. All these provide new ideas forfurther research in mechanism of microalgal oil production.