Insulin-like growth factor I stimulates degradation of an mRNA transcript encoding the 14 kDa ubiquitin-conjugating enzyme.
Insulin-like growth factor I stimulates degradation of an mRNA transcript encoding the 14 kDa ubiquitin-conjugating enzyme.
复制标题
胰岛素样生长因子 I 刺激编码 14 kDa 泛素结合酶的 mRNA 转录物降解。
作者：
S. Wing;N. Bédard
禁食后，骨骼肌中泛素依赖性蛋白水解系统被激活，同时蛋白水解速率增加。编码 14 kDa 泛素结合酶 (E2(14K)) 的 mRNA 水平可以催化该途径中的第一个不可逆反应，其水平与蛋白水解速率平行上升和下降 [Wing 和 Banville (1994) Am.J.生理学。 267，E39-E48]，表明泛素与蛋白质的缀合是一个受调控的步骤。为了表征这种调节机制，我们研究了胰岛素、胰岛素样生长因子 I (IGF-I) 和 des(1-3) 胰岛素样生长因子 I (DES-IGF-I)（不结合 IGF 结合蛋白）对 L6 肌管中 E2(14K) mRNA 水平的影响。胰岛素抑制 E2(14K) mRNA 水平，IC50 为 4 x 10(-9) M，但对编码多聚泛素和蛋白酶体亚基 C2 和 C8 的 mRNA 没有影响，与 E2(14K) 一样，禁食后骨骼肌也会增加。 E2(14K) mRNA水平的降低对IGF-I更敏感，IC50约为。 5 x 10(-10) M。在这些细胞孵育12小时期间，IGF-I结合蛋白显着分泌到培养基中。 DES-IGF-I 显着降低了对这些结合蛋白的亲和力，可有效降低 E2(14K) mRNA 水平，IC50 为 3 x 10(-11) M。DES-IGF-I 不会改变 E2(14K) 基因的转录速率，但会提高 1.2 kb mRNA 转录物的降解速率。 1.2 kb 转录本的半衰期大约是 1.8 kb 转录本的三分之一，并且可以解释之前观察到的该转录本的更显着的调节。这表明 1.8 kb 转录物中额外的 3' 非编码序列赋予了稳定性。这些观察结果表明，IGF-I 是 E2(14K) 表达的重要调节因子，并首次证明该激素会刺激特定 mRNA 转录物的降解，同时总体 RNA 会积累。
Upon fasting, the ubiquitin-dependent proteolytic system is activated in skeletal muscle in parallel with the increases in rates of proteolysis. Levels of mRNA encoding the 14 kDa ubiquitin-conjugating enzyme (E2(14K)), which can catalyse the first irreversible reaction in this pathway, rise and fall in parallel with the rates of proteolysis [Wing and Banville (1994) Am.J. Physiol. 267, E39-E48], indicating that the conjugation of ubiquitin to proteins is a regulated step. To characterize the mechanisms of this regulation, we have examined the effects of insulin, insulin-like growth factor I (IGF-I) and des(1-3) insulin-like growth factor I (DES-IGF-I), which does not bind IGF-binding proteins, on E2(14K) mRNA levels in L6 myotubes. Insulin suppressed levels of E2(14K) mRNA with an IC50 of 4 x 10(-9) M, but had no effects on mRNAs encoding polyubiquitin and proteasome subunits C2 and C8, which, like E2(14K), also increase in skeletal muscle upon fasting. Reduction of E2(14K) mRNA levels was more sensitive to IGF-I with an IC50 of approx. 5 x 10(-10) M. During the incubation of these cells for 12 h there was significant secretion of IGF-I-binding proteins into the medium. DES-IGF-I, which has markedly reduced affinity for these binding proteins, was found to potently reduce E2(14K) mRNA levels with an IC50 of 3 x 10(-11) M. DES-IGF-I did not alter rates of transcription of the E2(14K) gene, but enhanced the rate of degradation of the 1.2 kb mRNA transcript. The half-life of the 1.2 kb transcript was approximately one-third that of the 1.8 kb transcript and can explain the more marked regulation of this transcript observed previously. This indicates that the additional 3' non-coding sequence in the 1.8 kb transcript confers stability. These observations suggest that IGF-I is an important regulator of E2(14K) expression and demonstrate, for the first time, stimulation of degradation of a specific mRNA transcript by this hormone, while overall RNA accumulates.