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. Author manuscript; available in PMC: 2014 Mar 21.
Published in final edited form as: Mol Cell. 2011 Jul 8;43(1):33–44. doi: 10.1016/j.molcel.2011.04.028

Figure 6. PEPCK1 degradation is controlled by glucose level and can be manipulated by control degradation related factors.

Figure 6

(A)-(C), results are average values of triplicate q-RT-PCR assays with SDs. (A) P300 transcription is unregulated by glucose. P300 mRNA levels of 293T cells cultured under different glucose concentrations were determined. (B) Sirt2 transcription is down regulated by glucose. Sirt2 mRNA levels of 293T cells cultured under different glucose concentrations were determined. (C) Fasting reduces Sirt2 transcription. Sirt2 mRNA levels of mice liver cells were determined. (D) PEPCK1 ubiquitination response to glucose concentration. Flag-tagged PEPCK1 co-expressed with HA tagged ubiquitin in HEK293T cells maintained under various glucose concentrations were purified by IP and ubiquitination of purified proteins were determined by anti-HA antibody. (E)-(H), Secreted glucose levels of cells maintained in glucose free medium under conditions as indicated. Shown are average values of triplicate measurements with SDs. (E) Overexpression of UBR5-C decreases glucose production. HEK293T cells and HEK293T cells overexpressing UBR5-C and UBR5-C-C/A were analyzed. (F) UBR5 knockdown increases glucose production. Chang’s cells and Chang’s cells with UBR5 knocked down by siRNA were analyzed. (G) P300 knockdown increases glucose production. HEK293T cells and HEK293T cells with P300 knocked down by siRNA were analyzed. (H) Sirt2 knockdown decreases glucose production. HEK293T cells and HEK293T cells with Sirt2 knocked down by siRNA were analyzed. (I) Sirt2 knockdown decrease PEPCK1 and gluconeogenic rate in mice. Mice were tail vein injected by Sirt2 shRNA. Liver PEPCK1 level and blood glucose concentrations were measured. Shown are representative PEPCK1 protein levels and average blood concentrations with SD (n=4). See also Figure S4.