[Show abstract][Hide abstract] ABSTRACT: Pancreatic and duodenal homeobox 1 (PDX1) regulates pancreatic development and mature beta-cell function. We demonstrate by mass spectrometry that serine residue at position 269 in the C-terminal domain of PDX1 is phosphorylated in beta-cells. Besides we show that the degree of phosphorylation, assessed with a phospho-Ser-269-specific antibody, is decreased by elevated glucose concentrations in both MIN6 beta-cells and primary mouse pancreatic islets. Homeodomain interacting protein kinase 2 (HIPK2) phosphorylates PDX1 in vitro; phosphate incorporation substantially decreases in PDX1 S269A mutant. Silencing of HIPK2 led to a 51+/-0.2% decrease in Ser-269 phosphorylation in MIN6 beta-cells. Mutation of Ser-269 to phosphomimetic residue glutamic acid (S269E) or de-phosphomimetic residue alanine (S269A) exerted no effect on PDX1 half-life. Instead, PDX1 S269E mutant displayed abnormal changes in subnuclear localization in response to high glucose. Our results suggest that HIPK2-mediated phosphorylation of PDX1 at Ser-269 might be a regulatory mechanism connecting signals generated by changes in extracellular glucose concentration to downstream effectors via changes in subnuclear localization of PDX1, thereby influencing islet cell differentiation and function.
Biochemical and Biophysical Research Communications 08/2010; 399(2):155-61. DOI:10.1016/j.bbrc.2010.07.035 · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mammalian mitochondria contain about 1100 proteins, nearly 300 of which are uncharacterized. Given the well-established role of mitochondrial defects in human disease, functional characterization of these proteins may shed new light on disease mechanisms. Starting with yeast as a model system, we investigated an uncharacterized but highly conserved mitochondrial protein (named here Sdh5). Both yeast and human Sdh5 interact with the catalytic subunit of the succinate dehydrogenase (SDH) complex, a component of both the electron transport chain and the tricarboxylic acid cycle. Sdh5 is required for SDH-dependent respiration and for Sdh1 flavination (incorporation of the flavin adenine dinucleotide cofactor). Germline loss-of-function mutations in the human SDH5 gene, located on chromosome 11q13.1, segregate with disease in a family with hereditary paraganglioma, a neuroendocrine tumor previously linked to mutations in genes encoding SDH subunits. Thus, a mitochondrial proteomics analysis in yeast has led to the discovery of a human tumor susceptibility gene.
[Show abstract][Hide abstract] ABSTRACT: Metabolic disorders, such as diabetes and obesity, are fundamentally caused by cellular energy imbalance and dysregulation. Therefore, understanding the regulation of cellular fuel and energy metabolism is of great importance to develop effective therapies for metabolic disease. The cellular nutrient and energy sensors, AMPK and TOR, play a key role in maintaining cellular energy homeostasis. Like AMPK and TOR, PAS kinase (PASK) is also a nutrient responsive protein kinase. In yeast, PAS kinase phosphorylates the enzyme Ugp1 and thereby shifts glucose partitioning toward cell wall glucan synthesis at the expense of glycogen synthesis. Consistent with this function, yeast PAS kinase is activated by both cell integrity stress and growth in non-fermentative carbon sources. PASK is also important for proper regulation of glucose metabolism in mammals at both the hormonal and cellular level. In cultured pancreatic beta-cells, PASK is activated by elevated glucose concentrations and is required for glucose-stimulated transcription of the insulin gene. PASK knockdown in cultured myoblasts causes increased glucose oxidation and elevated cellular ATP levels. Mice lacking PASK exhibit increased metabolic rate and resistance to diet-induced obesity. Interestingly, PGC-1 expression and AMPK and TOR activity were not affected in PASK deficient mice, suggesting PASK may exert its metabolic effects through a new mechanism. We propose that PASK plays a significant role in nutrient sensing, metabolic regulation, and energy homeostasis, and is a potential therapeutic target for metabolic disease.
International Union of Biochemistry and Molecular Biology Life 04/2008; 60(4):204-9. DOI:10.1002/iub.32 · 3.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The metabolic syndrome, a complex set of phenotypes typically associated with obesity and diabetes, is an increasing threat to global public health. Fundamentally, the metabolic syndrome is caused by a failure to properly sense and respond to cellular metabolic cues. We studied the role of the cellular metabolic sensor PAS kinase (PASK) in the pathogenesis of metabolic disease by using PASK(-/-) mice. We identified tissue-specific metabolic phenotypes caused by PASK deletion consistent with its role as a metabolic sensor. Specifically, PASK(-/-) mice exhibited impaired glucose-stimulated insulin secretion in pancreatic beta-cells, altered triglyceride storage in liver, and increased metabolic rate in skeletal muscle. Further, PASK deletion caused nearly complete protection from the deleterious effects of a high-fat diet including obesity and insulin resistance. We also demonstrate that these cellular effects, increased rate of oxidative metabolism and ATP production, occur in cultured cells. We therefore hypothesize that PASK acts in a cell-autonomous manner to maintain cellular energy homeostasis and is a potential therapeutic target for metabolic disease.
Proceedings of the National Academy of Sciences 10/2007; 104(39):15466-71. DOI:10.1073/pnas.0705407104 · 9.67 Impact Factor