Sterol regulatory element binding protein-1a transactivates 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene promoter

6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB)catalyzes the synthesis and degradation of fructose-2,6-bisphosphate, a key modulator of glycolysis-gluconeogenesis. To gain insight into the molecular mechanism behind hormonal and nutritional regulation of PFKFB expression, we have cloned and characterized the proximal promoter region of the liver isoform of PFKFB (PFKFB1) from gilthead sea bream (Sparus aurata). Transient transfection of HepG2 cells with deleted gene promoter constructs and electrophoretic mobility shift assays allowed us to identify a sterol regulatory element (SRE) to which SRE binding protein-1a (SREBP-1a)binds and transactivates PFKFB1 gene transcription. Mutating the SRE box abolished SREBP-1a binding and transactivation. The in vivo binding of SREBP-1a to the SRE box in the S. aurata PFKFB1 promoter was confirmed by chromatin immunoprecipitation assays. There is a great deal of evidence for a postprandial rise of PFKB1 mRNA levels in fish and rats. Consistently, starved-to-fed transition and treatment with glucose or insulin increased SREBP-1 immunodetectable levels, SREBP-1 association to PFKFB1 promoter, and PFKFB1 mRNA levels in the piscine liver. Our findings demonstrate involvement of SREBP-1a in the transcriptional activation of PFKFB1, and we conclude that SREBP-1a may exert a key role mediating postprandial activation of PFKFB1 transcription

Impairment of Liver GH Receptor Signaling by Fasting

Fasting causes a state of GH resistance responsible for low circulating IGF-I levels. To investigate whether this resistance may result from alterations in the GH signaling pathway, we determined the effects of fasting on the GH transduction pathway in rat liver. Forty-eight-hour fasted or fed male rats were injected with recombinant rat GH via the portal vein. Liver was removed 0 and 15 min after injection. Although GH stimulated Janus kinase 2 (JAK2) phosphorylation in all animals, this was severely blunted in fasted animals. Similarly, the phosphorylation of the GH receptor, although observed in both fasted and fed rats after GH injection, was markedly reduced in fasted rats. A rapid signal transducer and activator of transcription 5 (STAT5) tyrosine phosphorylation was also induced in the liver of fed animals in response to GH. In contrast, in fasted rats only a slight phosphorylated STAT5 signal was observed. The inhibitory effect of fasting on these GH signaling molecules occurred without changes in their protein content. Furthermore, the impairment of the JAK-STAT pathway in fasted animals was associated with increased liver suppressor of cytokine signaling 3 mRNA levels. Although glucocorticoids, which are increased by fasting, may cause GH resistance, adrenalectomy failed to prevent alterations in the JAK-STAT pathway caused by fasting. In conclusion, the GH resistance induced by fasting is associated with impairment of the JAK-STAT signaling pathway. This might contribute to the decrease in liver IGF-I production observed in fasting