Smas C M, Chen L, Zhao L, Latasa M J, Sul H S
Department of Nutritional Sciences, University of California, Berkeley, California 94720, USA.
J Biol Chem. 1999 Apr 30;274(18):12632-41. doi: 10.1074/jbc.274.18.12632.
Pref-1 is an epidermal growth factor-like domain-containing transmembrane protein that is cleaved to generate a soluble factor. It is abundant in 3T3-L1 preadipocytes but absent in mature adipocytes. Constitutive expression of pref-1 or the addition of its ectodomain inhibits adipogenesis. We find that the pref-1 gene is an early target of dexamethasone, a component of the dexamethasone/methylisobutylxanthine differentiation mixture used routinely for adipoconversion. The time course of the decrease in pref-1 mRNA by dexamethasone reflected the pref-1 mRNA half-life determined by actinomycin D treatment. Nuclear run-on assays showed that dexamethasone attenuates pref-1 transcription. We demonstrate a correlation between pref-1 down-regulation and adipoconversion by varying the time period and concentration of dexamethasone. Increasing the dexamethasone treatment from 2 to 4 days resulted in a time-dependent pref-1 down-regulation and increased differentiation as measured by adipocyte marker mRNAs. The dexamethasone concentration between 1 and 10 nM showed a dose-dependent decrease in pref-1 mRNA and an enhancement of adipogenesis. To test the hypothesis that dexamethasone initiation of adipoconversion may be via down-regulation of pref-1, we lowered endogenous pref-1 mRNA levels by stably transfecting 3T3-L1 preadipocytes with antisense pref-1. At 1 microM, antisense cells had enhanced adipose conversion; a similar degree of differentiation occurred with 2 nM dexamethasone, a concentration that does not support differentiation of control 3T3-L1 cells. We conclude that dexamethasone-mediated repression of pref-1 contributes to the mechanisms whereby glucocorticoids promote adipogenesis.
Pref-1是一种含有表皮生长因子样结构域的跨膜蛋白，可被切割产生一种可溶性因子。它在3T3-L1前脂肪细胞中大量存在，但在成熟脂肪细胞中不存在。Pref-1的组成型表达或其胞外结构域的添加会抑制脂肪生成。我们发现pref-1基因是地塞米松的早期靶点，地塞米松是常用于脂肪转化的地塞米松/甲基异丁基黄嘌呤分化混合物的成分之一。地塞米松使pref-1 mRNA减少的时间进程反映了放线菌素D处理所确定的pref-1 mRNA半衰期。核转录分析表明地塞米松减弱了pref-1的转录。我们通过改变地塞米松的处理时间和浓度，证明了pref-1下调与脂肪转化之间的相关性。将地塞米松处理时间从2天增加到4天，导致pref-1随时间依赖性下调，并且通过脂肪细胞标志物mRNA测量的分化增加。1至10 nM之间的地塞米松浓度显示出pref-1 mRNA的剂量依赖性降低和脂肪生成的增强。为了检验地塞米松启动脂肪转化可能是通过下调pref-1这一假设，我们用反义pref-1稳定转染3T3-L1前脂肪细胞，降低内源性pref-1 mRNA水平。在1 microM时，反义细胞的脂肪转化增强；2 nM地塞米松也出现了类似程度的分化，而该浓度不支持对照3T3-L1细胞的分化。我们得出结论，地塞米松介导的pref-1抑制作用有助于糖皮质激素促进脂肪生成的机制。
