Internalization of the opioid growth factor, [Met5]-enkephalin, is dependent on clathrin-mediated endocytosis for downregulation of cell proliferation
Internalization of the opioid growth factor, [Met5]-enkephalin, is dependent on clathrin-mediated endocytosis for downregulation of cell proliferation
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DOI：
10.1152/ajpregu.00318.2010
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发表时间：
2010-09-01
影响因子：
2.8
通讯作者：
Zagon, Ian S.
中科院分区：
文献类型：
--
作者：
Cheng, Fan;McLaughlin, Patricia J.;Zagon, Ian S.
Cheng F，McLaughlin PJ，Banks WA，Zagon IS.阿片生长因子[Met（5）]-脑啡肽的内化依赖于网格蛋白介导的内吞作用，以下调细胞增殖。Am J Physiol Regul Integr Comp Physiol 299：R774-R785，2010年。首次发表于2010年6月30日; doi：10.1152/ajpregu.00318.2010。阿片样生长因子（OGF; [Met（5）]-脑啡肽）是一种组成性表达的具有张力活性的抑制性肽，与OGF受体（OGFr）相互作用，形成体内稳态的内源性生长调节途径。在动物和临床研究中，OGF-OGFr介导的扩增抑制发育、肿瘤形成、血管生成和免疫。OGF-OGFr轴的破坏加速细胞增殖，并且在伤口修复中特别重要。为了研究OGF如何进入细胞，用5，6-四甲基罗丹明OGF（RhoOGF）标记OGF以研究其在活细胞中的摄取。用RhoOGF孵育的非洲绿色猴肾细胞（COS-7）显示出温度依赖性的进入过程，在37 ℃下内化，但在4 ℃下不内化。初始暴露15 min后，在细胞质中检测到RhoOGF，30 min内在细胞质和细胞核中均观察到RhoOGF，并在细胞中保留长达5 h。过量100倍的OGF或阿片拮抗剂纳洛酮，而不是其他阿片配体（一些选择性的经典阿片受体），显着减少RhoOGF进入细胞。RhoOGF是功能性的，因为与RhoOGF（10（-5）至10（-8）M）孵育的细胞中的DNA合成降低24- 36%，并且与用未标记的OGF处理的细胞相当（降低26-39%）。OGF内化依赖于网格蛋白介导的内吞作用，加入网格蛋白siRNA减少RhoOGF的摄取并上调DNA合成。RhoOGF网格蛋白介导的内吞作用与内体或高尔基体途径无关。两者合计，这些结果表明，OGF进入细胞的主动运输在一个饱和的方式，需要网格蛋白介导的内吞作用。
Cheng F, McLaughlin PJ, Banks WA, Zagon IS. Internalization of the opioid growth factor, [Met(5)]-enkephalin, is dependent on clathrin-mediated endocytosis for downregulation of cell proliferation. Am J Physiol Regul Integr Comp Physiol 299: R774-R785, 2010. First published June 30, 2010; doi:10.1152/ajpregu.00318.2010.-The opioid growth factor (OGF; [Met(5)]-enkephalin), a constitutively expressed and tonically active inhibitory peptide, interacts with the OGF receptor (OGFr) to form an endogenous growth-regulating pathway in homeostasis. Amplification of OGF-OGFr interfacing in animal and clinical studies depresses development, neoplasia, angiogenesis, and immunity. Disruption of the OGF-OGFr axis accelerates cell proliferation and has been particularly important in wound repair. To investigate how OGF enters cells, OGF was labeled with 5,6-tetramethylrhodamine OGF (RhoOGF) to study its uptake in live cells. African green monkey kidney cells (COS-7) incubated with RhoOGF exhibited a temperature-dependent course of entry, being internalized at 37 C but not at 4 C. RhoOGF was detected in the cytoplasm 15 min after initial exposure, observed in both cytoplasm and nucleus within 30 min, and remained in the cells for as long as 5 h. A 100-fold excess of OGF or the opioid antagonist naltrexone, but not other opioid ligands (some selective for classic opioid receptors), markedly reduced entry of RhoOGF into cells. RhoOGF was functional because DNA synthesis in cells incubated with RhoOGF (10(-5) to 10(-8) M) was decreased 24-36%, and was comparable to cells treated with unlabeled OGF (reductions of 26-39%). OGF internalization was dependent on clathrin-mediated endocytosis, with addition of clathrin siRNA diminishing the uptake of RhoOGF and upregulating DNA synthesis. RhoOGF clathrin-mediated endocytosis was unrelated to endosomal or Golgi pathways. Taken together, these results suggest that OGF enters cells by active transport in a saturable manner that requires clathrin-mediated endocytosis.