The Mammalian INDY Homolog Is Induced by CREB in a Rat Model of Type 2 Diabetes

Abstract

Reduced expression of the INDY (I'm not dead yet) tricarboxylate carrier increased the life span in different species by mechanisms akin to caloric restriction. Mammalian INDY homolog (mIndy, SLC13A5) gene expression seems to be regulated by hormonal and/or nutritional factors. The underlying mechanisms are still unknown. The current study revealed that mIndy expression and [14C]-citrate uptake was induced by physiological concentrations of glucagon via a cAMP-dependent and cAMP-responsive element–binding protein (CREB)–dependent mechanism in primary rat hepatocytes. The promoter sequence of mIndy located upstream of the most frequent transcription start site was determined by 5′-rapid amplification of cDNA ends. In silico analysis identified a CREB-binding site within this promoter fragment of mIndy. Functional relevance for the CREB-binding site was demonstrated with reporter gene constructs that were induced by CREB activation when under the control of a fragment of a wild-type promoter, whereas promoter activity was lost after site-directed mutagenesis of the CREB-binding site. Moreover, CREB binding to this promoter element was confirmed by chromatin immunoprecipitation in rat liver. In vivo studies revealed that mIndy was induced in livers of fasted as well as in high-fat-diet–streptozotocin diabetic rats, in which CREB is constitutively activated. mIndy induction was completely prevented when CREB was depleted in these rats by antisense oligonucleotides. Together, these data suggest that mIndy is a CREB-dependent glucagon target gene that is induced in fasting and in type 2 diabetes. Increased mIndy expression might contribute to the metabolic consequences of diabetes in the liver.

Differential response of the natriuretic peptide system to weight loss and exercise in overweight or obese patients

“…mRproANP was detected in plasma using a sandwich immunoassay in a chemiluminescence coated tube format (BRAHMS AG, Hennigsdorf/Berlin, Germany) as described previously [13]. We measured NPR-A and NPR-C gene expression in abdominal subcutaneous adipose tissue samples obtained by needle biopsies as described [14]. Quantities were interpolated from diluted standard curves, which account for reaction efficiencies [14].…”

“…We measured NPR-A and NPR-C gene expression in abdominal subcutaneous adipose tissue samples obtained by needle biopsies as described [14]. Quantities were interpolated from diluted standard curves, which account for reaction efficiencies [14]. Target genes were normalized to relative expression levels of hypoxanthine-guanine phosphoribosyltransferase (HPRT).…”

The Role of Indy in Metabolic Regulation

“…The promoter sequence of mINDY was identified including a CREB binding site within this fragment, identifying mINDY as a CREB-dependent glucagon target gene, which is induced in the short term fasting status and type-2 diabetes [62]. …”

Inhibition of citrate cotransporter Slc13a5/mINDY by RNAi improves hepatic insulin sensitivity and prevents diet-induced non-alcoholic fatty liver disease in mice

“…Genetic whole-body deletion of mINDY in mice also led to striking similarities to caloric restriction and protected mice from high-fat diet (HFD)-induced obesity, steatosis hepatis, and IR by increasing energy expenditure, improving hepatic mitochondrial biogenesis, enhancing hepatic FA oxidation, and reducing hepatic lipogenesis [16]. In primary rat hepatocytes, mINDY expression and citrate uptake could be induced by physiological concentrations of glucagon via a cAMP-dependent and cAMP-responsive element-binding protein (CREB)-dependent mechanism at a confirmed CREB-binding site within the mINDY promoter [23]. In line with these observations, the siRNA-mediated knockdown of mINDY in human HepG2 cells reduced their lipid content [24].…”