The Err To Promote Glucokinase Expression Of Pgc-1¦Â Regulate The Preliminary Study Of The Mechanism Of Transcription Of Alas1 And Mitochondrial Gene Promoter

The orphan nuclear receptor estrogen-related receptors(ERR a and ERRγ) are activated by the transcriptional coactivator peroxisome proliferator-activated receptorγ(PPARγ) coactivator 1α(PGC-1α), a critical regulator of cellular energy metabolism. Glucokinase plays a key role in the regulation of glucose utilization in liver and its expression is strongly enhanced by insulin. We report here that the regulatory sequences of the GK gene harbor a functional ERRαand ERRγbinding site and the co-stimulating effects of ERRs and PGC-1αon GK gene expression in the rat primary hepatocytes, suggesting a novel role for ERRs in controlling glucose metabolism. The co-stimulating effect was inhibited when the GK promoter fragment was mutated at the ERRE. In contrast to the dramatic effects of ERRs and PGC-1αon full length GK, they had only a small effect on the truncated GK construct that lacked the ERRE. By using a synthetic inhibitor of ERRα, the inhibitory result revealed that ERRαplay a key role in insulin stimulating GK expression. Furthermore, ERRαand ERRγincreased endogenous GK mRNA, protein and GK activity. Our data demonstrate that ERRs can directly activate GK expression in liver and may contribute to improving glucose homeostasis in type 2 diabetes. PGC-1βis a recently identified transcriptional coactivator closely related to PGC-1αwhose biological activities are largely unknown. Like PGC-1α, PGC-1βstrongly activates mitochondrial biogenesis and cellular respiration in differentiated myotubes. Heme is an essential component of numerous hemoproteins with functions including oxygen transport, energy metabolism.5-aminolevulinate synthase (ALAS1) is the rate-limiting enzyme in heme biosynthesis which provide heme for cytochromes in the respiratory chain. By 5'promoter deletion assay, we found PGC-1βcould activate ALAS 1 promoter that depended on NRF 1 for full activity. The ALAS 1 promoter has two functional NRF-1 recognition sites that are essential for basal promoter activity.When either of the two NRF-1 elements was mutated, about 60-70% of promoter activity was lost; when both were mutated, full activity was diminished by approximately 90%. Furthermore, expression of a dominant negative NRF-1 inhibited the PGC-1βmediated increase in ALAS1 gene. PGC-1βcould also bind and co-activate NRF-1 and augment transcriptional activation of mitochondrial related gene. To determine whether NRF-1 is required for the expression of endogenous PGC-1βtargets, we used an adenovial RNAi vector (Ad-RNAi) toward NRF-1. Treatment of C2C12 myotubes with this Ad-RNAi reduces endogenous NRF-1 protein by approximately 70%. Ad-PGC-1βexpression in C2C12 myotubes stimulated mRNA abundance of several mitochondrial related gene such as ALAS-1, Cytc, ATPase, Tfb2m, Tfam, COX7b, MRPL46. The induction of all these genes in response to PGC-1β, however, is greatly impaired in the cells infected with Ad-RNAi compared to the control GFP. These results suggest that PGC-1βplay an important role in controlling mitochondrial oxidative energy metabolism.