Fig. 1: The protein–protein interaction of Arabidopsis glycolytic enzymes.

a The protein interactions calculated fold change in the heatmap. The novel complex of phosphoglycerate mutase 1–enolase–PYK4 complex could be detected. The star mark was represented fold change more than four times compared with GFP control. b Sublocation of all the glycolysis according to the SUBA4 database mitochondria proteomics data and GFP signal. c NE-TPT/PGAM1-CE. d NE-TPT/PGAM2-CE. e Enolase-NE/PGAM1-CE. f Enolase-NE/PGAM2-CE. g Enolase-NE/PYK4-CE. h VDAC1-NE/PYK4-CE. i VDAC3-NE/PYK4-CE. Confirmation of phosphoglycerate mutase 1–enolase–PYK4 complex with membrane protein (chloroplast TPT and mitochondria VDAC) by bimolecular fluorescent complementation (BiFC) assay. Interactions among chloroplast and mitochondria membrane protein and phosphoglycerate mutase 1–enolase–PK4 complex were further tested by BiFC with transient expression of tagged proteins in Arabidopsis mesophyll protoplasts. NE is the N-terminal of the split-mCitrine, CE is the C-terminal of the split-mCitrine. The panels from the left side show the BiFC fluorescence, fluorescence from bright-field image, MitoTracker Red staining, autofluorescence, blank, and the merged image of all of those, respectively. j Confirmation of protein–protein interaction by split Renilla luciferase assay. Interactions among chloroplast and mitochondria membrane protein and phosphoglycerate mutase–enolase–pyruvate kinase complex were further confirmed by split Renilla luciferase with transient expression of tagged proteins in Arabidopsis mesophyll protoplasts. One-way ANOVA analysis by enolase/GFP as a negative control (*P < 0.05, SD). PGM phosphoglucomutase, HXK hexokinase, PGI phosphoglucose isomerase, PFK phosphofructokinase, ALD aldolase, TPI tripsephosphate isomerase, GAPDH glyceraldehyde phosphate dehydrogenase, PGK phosphoglycerate kinase, ENO enolase, PYK pyruvate kinase, PGAM phosphoglycerate mutase, TPT triose phosphate translocator, VDAC voltage-dependent anion channel.