ENDOGENOUS ORIGIN OF DEFECTIVE RETROVIRUS-LIKE PARTICLES FROM A RECOMBINANT CHINESE-HAMSTER OVARY CELL-LINE
ENDOGENOUS ORIGIN OF DEFECTIVE RETROVIRUS-LIKE PARTICLES FROM A RECOMBINANT CHINESE-HAMSTER OVARY CELL-LINE
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DOI：
10.1016/0042-6822(91)90496-x
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发表时间：
1991-03-01
期刊：
影响因子：
3.7
通讯作者：
DINOWITZ, M
中科院分区：
文献类型：
--
作者：
ANDERSON, KP;LOW, MAL;DINOWITZ, M
中国仓鼠卵巢（CHO）细胞中出芽的C型和胞浆内A型颗粒的存在已得到充分证实。然而，广泛的筛查未能发现任何传染性的证据。连续流超离心已被用于从重组CHO细胞亚克隆的培养液中浓缩细胞外颗粒以用于分子表征。颗粒表现出逆转录酶活性，并与哺乳动物C型逆转录病毒结构蛋白相关，以逆转录病毒的特征密度在蔗糖梯度中形成条带。通过电子显微镜检查梯度纯化的颗粒显示与其他逆转录病毒相似的形态和大小。双梯度纯化的颗粒含有与鼠白血病病毒探针杂交的RNA，和内源性中国仓鼠脑池内A颗粒元件。从纯化的颗粒分离的cDNA克隆的DNA序列分析揭示了核酸内切酶阅读框的多次中断，为观察到的颗粒的非感染性性质提供了一种可能的解释。纯化颗粒中以RNA形式存在的序列也以保守、重复的前病毒序列形式存在于检查的所有CHO细胞系的基因组DNA和中国仓鼠肝DNA中。因此，观察到的颗粒可能是中国仓鼠生殖系中存在的内源性逆转录病毒样成分的产物。
The presence of budding C-type and intracytoplasmic A-type particles in Chinese hamster ovary (CHO) cells is well documented. However, extensive screening has failed to detect any evidence of infectivity. Continuous-flow ultracentrifugation has been used to concentrate extracellular particles from culture fluid of a recombinant CHO cell subclone for molecular characterization. Particles exhibiting reverse transcriptase activity and associated with mammalian C-type retrovirus structural proteins banded in sucrose gradients at a density characteristic of retroviruses. Examination of gradient-purified particles by electron microscopy revealed morphology and size similar to other retroviruses. Double-gradient-purified particles contained RNA which hybridized to probes for murine leukemia virus, and endogenous Chinese hamster intracisternal A-particle elements. DNA sequence analysis of a cDNA clone isolated from purified particles revealed multiple interruptions of the endonuclease reading frame, providing one possible explanation for the noninfectious nature of the observed particles. Sequences present as RNA in purified particles were also present as conserved, repetitive, provirus sequences in genomic DNA of all CHO cell lines examined and in Chinese hamster liver DNA. The observed particles are therefore likely to be the products of endogenous retroviruslike elements present in the germline of Chinese hamsters.