Abstract
Krüppel-like factor 14 (KLF14) is a member of the Cys2/His2 zinc-finger DNA-binding proteins. Despite strong evidence showing that a polymorphism in the Klf14 gene is closely linked to the development of type 2 diabetes, the physiological and metabolic functions of KLF14 still remain unclear. In the present study, we investigated the role of KLF14 in the regulation of hepatic gluconeogenesis. Adenoviruses expressing KLF14 (Ad-Klf14) or KLF14-specific shRNAs (Ad-shKlf14) were injected into normal C57BL/6J, db/db diabetic, or high-fat diet-induced obese (DIO) mice. Gene expression, hepatic glucose production, glucose tolerance, and insulin resistance were tested in C57BL/6J, db/db, and DIO mice and primary hepatocytes. Our results demonstrate that KLF14 expression is induced in the livers of normal C57BL/6J mice upon fasting and significantly up-regulated in the livers of db/db mice, suggesting a physiological link between KLF14 and gluconeogenesis. Adenovirus-mediated overexpression of KLF14 in primary hepatocytes increased both the mRNA and protein levels of peroxisome proliferator-activated receptor-γ coactivator 1α (Pgc-1α, also known as Ppargc1a), thereby stimulating cellular glucose production. Conversely, knockdown of KLF14 expression led to a reduction in PGC-1α, subsequently inhibiting glucose output in primary hepatocytes. Finally, forced expression of KLF14 in the livers of normal mice increased the plasma glucose levels and impaired glucose tolerance; in contrast, KLF14 knockdown in diabetic mouse livers improved glucose tolerance. Taken together, our data strongly indicate that KLF14 modulates hepatic gluconeogenesis.
Highlights
Krüppel-like factor 14 (KLF14) is a member of the Cys2/His2 zinc-finger DNA-binding proteins
Genetic evidence from various studies has shown that Klf14 is closely related to metabolic diseases, including neonatal/juvenile diabetes and metabolic syndrome with obesity-associated insulin resistance, leading to adult Type 2 diabetes (T2D) [15, 16, 24]
The results demonstrated that overnight fasting led to a significant induction in both Klf14 mRNA and protein levels in the liver and that the induction was restored upon refeeding (Fig. 1A)
Summary
T2D, type 2 diabetes; Ad-gfp, adenovirus containing green fluorescent protein; Ad-Klf, adenovirus expressing KLF14; Ad-shCtrl, control adenovirus expressing short-hairpin RNA against luciferase; Ad-shKlf, adenovirus expressing short hairpin RNA against KLF14; DEX, dexamethasone; DIO, diet-induced obese; FSK, forskolin; GTT, glucose tolerance test; ITT, insulin tolerance test; PEPCK, phosphoenolpyruvate carboxykinase; PTT, pyruvate tolerance test; TG, triglyceride; FFA, free fatty acid; HFD, high-fat diet; ALT, alanine aminotransferase; AST, aspartate aminotransferase; qPCR, quantitative PCR. PGC-1␣ expression is tightly regulated in response to low systemic glucose levels. PGC-1␣ transactivates gluconeogenic gene expression by coactivating a variety of transcription factors, including hepatic nuclear factor-4␣ (HNF4␣), glucocorticoid receptor, forkhead box O1 (FOXO1), and cAMP-responsive element–binding protein [22]. These transcription factors bind to the promoter regions of genes that encode key gluconeogenic enzymes, such as phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase [23]. Our results indicated that adenovirus-mediated overexpression of KLF14 in mouse primary hepatocytes increased the master regulator of hepatic gluconeogenesis, PGC-1␣, as well as PGC-1␣ target gene expression. The present study demonstrated that the KLF14 is a critical physiological regulator of hepatic gluconeogenesis
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