Pooled CRISPR/Cas9 reveals redundant roles of plastidial phosphoglycerate kinases in carbon fixation and metabolism. (4th April 2019)

• Record Type: Journal Article
• Authors: Li, Ruizi; Qiu, Zhimin; Wang, Xiaoguo; Gong, Pingping; Xu, Qinzhen; Yu, Qing‐bo; Guan, Yuefeng
• Is Part Of: Plant journal. Volume 98:Number 6(2019)
• Page Start: 1078
• Page End: 1089
• Publication Date: 2019-04-04
• Subjects: Phosphoglycerate kinase -- glycolysis -- photosynthesis -- galactoglycerolipids -- carbon metabolism -- carbon fixation; Plant molecular biology -- Periodicals; Plant cells and tissues -- Periodicals; Botany -- Periodicals; 580
• DOI: 10.1111/tpj.14303
• Languages: English
• ISSNs: 0960-7412

Abstract

Summary: Phosphoglycerate kinase (PGK) is a highly conserved reversible enzyme that participates in both glycolysis and photosynthesis. In Arabidopsis thaliana, one cytosolic PGK (PGKc) and two plastidial PGKs (PGKp) are known. It remains debatable whether the two PGKp isozymes are functionally redundant or specialized in plastidial carbon metabolism and fixation. Here, using a pooled clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9) strategy, we found that plants with single mutations in pgkp1 or pgkp2 were not significantly affected, whereas a pgkp1pgkp2 double mutation was lethal due to retarded carbon fixation, suggesting that PGKp isozymes play redundant functional roles. Metabolomic analysis demonstrated that the sugar‐deficient pgkp1pgkp2 double mutation was partially complemented by exogenous sugar, although respiration intermediates were not rescued. Chloroplast development was defective in pgkp1pgkp2, due to a deficiency in glycolysis‐dependent galactoglycerolipid biosynthesis. Ectopic expression of a plastid targeting PGKc did not reverse the pgkp1pgkp2 double‐mutant phenotypes. Therefore, PGKp1 and PGKp2 play redundant roles in carbon fixation and metabolism, whereas the molecular function of PGKc is more divergent. Our study demonstrated the functional conservation and divergence of glycolytic enzymes. Significance Statement: There have been debates on whether different plastidial PGK isoforms play specialized or redundant roles in plastidial glycolysis and the Calvin cycle. Utilizing CRISPR/Cas9 and biochemical, metabolomic and lipidomic approaches, we found that Arabidopsis PGKp isozymes serve redundant, but essential, functions in carbon fixation, metabolism and galactoglycerolipids biosynthesis in chloroplasts.