Mihara A, Fujiwara K, Sato S, Okabe T, Fujiyoshi N
In Vitro Cell Dev Biol. 1987 Apr;23(4):317-22. doi: 10.1007/BF02623717.
A human erythroleukemia cell line, K-562 T1, was adapted to a protein-free chemically defined medium; that is, the medium does not contain any proteins such as exogenous hormones, growth factors, serum and serum albumin. The K-562 T1 cells which can proliferate in a protein-free medium are one of the model systems suitably supporting the autocrine hypothesis, which claims that cancer cells produce and respond to their own growth factors. The K-562 T1 cells were cultured in a protein-free medium at large scale and the growth factors were purified from the conditioned medium. It was found that K-562 T1 cells produce at least two growth factors; one is LGF-I (leukemia-derived growth factor-I) which can stimulate the proliferation of a wide range of human leukemia cell lines and the other is LGF-II (leukemia-derived growth factor-II), which can contribute to the growth of fibroblasts. LGF-I was purified using QAE-Sephadex, Bio Gel P-60 and Mono S FPLC. The purified protein was found to be homogenous by SDS-polyacrylamide gel electrophoresis and NH2-terminal sequence analysis. The molecular weight of LGF-I was 20,000 by SDS-polyacrylamide gel electrophoresis. The 30 NH2-terminal residues of LGF-I are the same as that of ubiquitin. Ubiquitin is a protein found in eukaryotic cells with molecular weight of 8,600. In the nucleus ubiquitin is conjugated to histone 2A to form the nuclear protein A24 which may play a role in regulation of chromatin structure, and in the cytoplasm is part of an ATP-dependent non-lysosomal proteolytic pathway. However, its physiological significance has not yet been fully resolved. Ubiquitin purified from bovine thymus did not show cell proliferating activity for any cells tested. The results suggest that LGF-I is a new autocrine growth factor with a molecular weight of 20,000 daltons, containing ubiquitin at the NH2-terminal end.
一种人红白血病细胞系K-562 T1被适应于无蛋白化学限定培养基；也就是说，该培养基不含任何蛋白质，如外源激素、生长因子、血清和血清白蛋白。能够在无蛋白培养基中增殖的K-562 T1细胞是适合支持自分泌假说的模型系统之一，该假说认为癌细胞产生并对自身生长因子作出反应。K-562 T1细胞在无蛋白培养基中大规模培养，并从条件培养基中纯化生长因子。发现K-562 T1细胞产生至少两种生长因子；一种是LGF-I（白血病衍生生长因子-I），它能刺激多种人白血病细胞系的增殖，另一种是LGF-II（白血病衍生生长因子-II），它能促进成纤维细胞的生长。LGF-I使用QAE-葡聚糖凝胶、Bio Gel P-60和Mono S FPLC进行纯化。通过SDS-聚丙烯酰胺凝胶电泳和NH2末端序列分析发现纯化的蛋白质是同质的。通过SDS-聚丙烯酰胺凝胶电泳，LGF-I的分子量为20,000。LGF-I的30个NH2末端残基与泛素的相同。泛素是一种在真核细胞中发现的蛋白质，分子量为8,600。在细胞核中，泛素与组蛋白2A结合形成核蛋白A24，其可能在染色质结构调节中起作用，在细胞质中是ATP依赖性非溶酶体蛋白水解途径的一部分。然而，其生理意义尚未完全阐明。从牛胸腺中纯化的泛素对任何测试细胞均未显示细胞增殖活性。结果表明，LGF-I是一种分子量为20,000道尔顿的新型自分泌生长因子，在NH2末端含有泛素。
