Analysis of the Glucose-Dependent Transcriptome in Murine Hypothalamic Cells

Abstract

We have previously shown that glucose activates CREB-regulated transcriptional co-activator-2 (CRTC-2) in murine, hypothalamic (mHypoA-2/10) cells. Thus, we now analysed the entire glucose-dependent transcriptome of mHypoA-2/10 cells by total RNA-seq. 831 genes were up- and 1390 genes down-regulated by at least 50 %. Signalling pathway analysis revealed activation of the cholesterol biosynthesis pathway by glucose. Accordingly, protein expression of both sterol regulatory element-binding proteins (SREBP) and total cholesterol levels were enhanced by glucose. Analysis of single genes involved in fundamental signalling processes suggested a significant impact of glucose. Thus, we chose ~100 genes and validated the effects of glucose on mRNA levels by qRT-PCR. We identified 15 genes with strong glucose-dependent mRNA expression. Among these genes were gnai1 to -3, adyc6, irs1, igfr1, hras and elk3. cAMP measurements revealed decreased basal and enhanced noradrenalin-induced cAMP levels at higher glucose concentrations. Serum-response element-dependent reporter assays indicated enhanced basal and insulin-like growth factor-induced activity at higher glucose levels. siRNA against CRTC-2 dampened the effects of glucose on cholesterol synthesis and IRS-1, SREBP-1, SREBP-2 or AC-6 protein expression. These findings could help to understand the functional consequences of physiologically occurring alterations of extracellular glucose concentrations as well as pathologically increased glucose levels.