Alteration of epidermal growth factor-dependent phosphorylation during rat liver regeneration.
Alteration of epidermal growth factor-dependent phosphorylation during rat liver regeneration.
复制标题
大鼠肝脏再生过程中表皮生长因子依赖性磷酸化的改变。
DOI：
10.1073/pnas.79.3.776
复制
发表时间：
1982
影响因子：
11.1
通讯作者：
Earp,HS
中科院分区：
文献类型：
--
作者：
Rubin,RA;O'Keefe,EJ;Earp,HS
表皮生长因子（EGF）刺激人类细胞系A-431中的膜蛋白磷酸化。已知的肝促有丝分裂作用的EGF和EGF受体的数量减少，发生在肝再生导致我们研究是否EGF依赖性蛋白激酶活性存在于大鼠肝脏和其活性是否改变后，部分肝切除术。肝膜，预孵育或没有EGF，磷酸化（0 ℃，15秒），并进行NaDodSO 4/聚丙烯酰胺凝胶电泳和放射自显影。在微粒体组分中，5-2000 ng/ml的EGF产生剂量相关的刺激，32 P掺入到单个170，000-道尔顿蛋白（p170）中。在质膜中，存在类似的EGF依赖性磷酸化，并且相对于微粒体组分显著富集。酸水解标记的微粒体组分，然后磷酸化氨基酸测定显示，EGF刺激32 P掺入磷酸化酪氨酸残基。部分肝切除术或假手术后36小时，从大鼠分离的微粒体组分中比较EGF依赖的p170磷酸化。在没有EGF的情况下，p170的体外标记是相似的。EGF刺激标记的p170在两组中，但反应明显减弱后，部分肝切除术。在EGF的存在下，再生肝脏微粒体组分中的p170标记仅为假手术大鼠膜中观察到的p170标记的47 +/- 6%（p <0.005）。肝再生过程中EGF依赖性磷酸化的减少导致125 I标记的EGF结合丧失。在部分肝切除术后，还观察到130，000-道尔顿蛋白的EGF非依赖性磷酸化的增加。这种磷蛋白的量的增加大致等于EGF刺激的p170磷酸化的损失。几个额外的蛋白质表现出增加磷酸化的膜部分肝切除大鼠。这些发现表明，在膜酪氨酸残基磷酸化的改变发生在体内调节生长。
Epidermal growth factor (EGF) stimulates membrane protein phosphorylation in a human cell line, A-431. The known hepatic mitogenic action of EGF and the reduction in EGF receptor number that occurs during liver regeneration led us to study whether EGF-dependent protein kinase activity was present in rat liver and whether its activity was altered after partial hepatectomy. Liver membranes, preincubated with or without EGF, were phosphorylated (0 degrees C, 15 sec) and subjected to NaDodSO4/polyacrylamide gel electrophoresis and autoradiography. In microsomal fractions, EGF at 5-2000 ng/ml produced a dose-related stimulation of 32P incorporation into a single 170,000-dalton protein (p170). In plasma membranes, a similar EGF-dependent phosphorylation was present and was substantially enriched relative to the microsomal fraction. Acid hydrolysis of labeled microsomal fraction followed by phosphoamino acid determination revealed that EGF stimulated 32P incorporation into phosphotyrosine residues. The EGF-dependent phosphorylation of p170 was compared in microsomal fractions isolated from rats 36 hr after partial hepatectomy or sham operation. In the absence of EGF, in vitro labeling of p170 was similar. EGF stimulated the labeling of p170 in both groups, but the response was clearly diminished after partial hepatectomy. In the presence of EGF, the labeling of p170 in microsomal fraction from regenerating livers was only 47 +/- 6% of that observed in membranes from sham-operated rats (p less than 0.005). Reduction of EGF-dependent phosphorylation during liver regeneration paralleled the loss of binding of 125I-labeled EGF. An increase in the EGF-independent phosphorylation of a 130,000-dalton protein was also observed after partial hepatectomy. The increase in the amount of this phosphoprotein was roughly equal to the loss of EGF-stimulated p170 phosphorylation. Several additional proteins showed increased phosphorylation in membranes from partially hepatectomized rats. These findings indicate that alterations in membrane tyrosine residue phosphorylation occur during regulated growth in vivo.