Fig 6. Growth of the ^RNAiPRODH.i mutant is rescued by α-ketoglutarate.

The efficiency of RNAi-mediated downregulation of PRODH expression in the tetracycline-induced (.i) or non induced (.ni) ^RNAiPRODH cell line, as well as in the parental cell line (WT) was determined by the PRODH activity assay (panel A) and ¹H-NMR quantification of end-products excreted from metabolism of proline and glucose in two independent experiments (panel B). Panel C shows growth curves of the ^RNAiPRODH.i cell line in glucose-depleted medium containing 2 mM proline, in the presence (colored circles) or absence (open circles) of 10 mM α-ketoglutarate, malate, succinate or proline. Cells were maintained in the exponential growth phase and cumulative cell numbers reflect normalization for dilution during cultivation. The effect of 10 mM succinate, malate, α-ketoglutarate or proline on growth of the parental and ^RNAiPRODH.i cell lines in glucose-depleted medium containing 2 mM proline, using the Alamar Blue assay described in Fig 4, is shown in panel D.