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Hirata M.,Tokyo University of Pharmacy and Life Science | Suzuki M.,Tokyo University of Pharmacy and Life Science | Ishii R.,Tokyo University of Pharmacy and Life Science | Satow R.,Tokyo University of Pharmacy and Life Science | And 8 more authors.
Diabetes | Year: 2011

OBJECTIVE - Regulation of obesity development is an important issue to prevent metabolic syndromes. Gene-disrupted mice of phospholipase Cδ1 (PLCδ1), a key enzyme of phosphoinositide turnover, seemed to show leanness. Here we examined whether and how PLCδ1 is involved in obesity. RESEARCH DESIGN AND METHODS - Weight gain, insulin sensitivity, and metabolic rate in PLCδ1-/- mice were compared with PLCδ1 +/- littermate mice on a high-fat diet. Thermogenic and adipogenetic potentials of PLCδ1-/- immortalized brown adipocytes and adipogenesis of PLCδ1-knockdown (KD) 3T3L1 cells, or PLCδ1 -/- white adipose tissue (WAT) stromal-vascular fraction (SVF) cells, were also investigated. RESULTS - PLCδ1-/- mice showed marked decreases in weight gain and mass of epididymal WAT and preserved insulin sensitivity compared with PLCδ1+/- mice on a high-fat diet. In addition, PLCδ1-/- mice have a higher metabolic rate such as higher oxygen consumption and heat production. When control immortalized brown adipocytes were treated with thermogenic inducers, expression of PLCδ1 was decreased and thermogenic gene uncoupling protein 1 (UCP1) was upregulated to a greater extent in PLCδ1-/- immortalized brown adipocytes. In contrast, ectopic expression of PLCδ1 in PLCδ1-/- brown adipocytes induced a decrease in UCP expression, indicating that PLCδ1 negatively regulates thermogenesis. Importantly, accumulation of lipid droplets was severely decreased when PLCδ1-KD 3T3L1 cells, or PLCδ1 -/- WAT SVF cells, were differentiated, whereas differentiation of PLCδ1-/- brown preadipocytes was promoted. CONCLUSIONS - PLCδ1 has essential roles in thermogenesis and adipogenesis and thereby contributes to the development of obesity. © 2011 by the American Diabetes Association.

Hashimoto Y.,Institute for Molecular and Cellular Regulation | Hashimoto Y.,Gunma University | Muramatsu K.,Institute for Molecular and Cellular Regulation | Muramatsu K.,Nara Institute of Science and Technology | And 9 more authors.
FASEB Journal | Year: 2012

The molecular mechanisms of neuronal morphology and synaptic vesicle transport have been largely elusive, and only a few of the molecules involved in these processes have been identified. Here, we developed a novel morphology-based gene trap method, which is theoretically applicable to all cell lines, to easily and rapidly identify the responsible genes. Using this method, we selected several genetrapped clones of rat pheochromocytoma PC12 cells, which displayed abnormal morphology and distribution of synaptic vesicle-like microvesicles (SLMVs). We identified several genes responsible for the phenotypes and analyzed three genes in more detail. The first gene was BTB/POZ domain-containing protein 9 (Btbd9), which is associated with restless legs syndrome. The second gene was cytokine receptor-like factor 3 (Crlf3), whose involvement in the nervous system remains unknown. The third gene was single-stranded DNA-binding protein 3 (Ssbp3), a gene known to regulate head morphogenesis. These results suggest that Btbd9, Crlf3, and Ssbp3 regulate neuronal morphology and the biogenesis/transport of synaptic vesicles. Because our novel morphology-based gene trap method is generally applicable, this method is promising for uncovering novel genes involved in the function of interest in any cell lines. © The Author(s).

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