Loh T P, Sievert L L, Scott R W
E. I. du Pont de Nemours and Company, Inc., Central Research and Development Department, Wilmington, Delaware 19898.
Mol Cell Biol. 1987 Oct;7(10):3775-84. doi: 10.1128/mcb.7.10.3775-3784.1987.
Embryonal carcinoma (EC) cells are nonpermissive for retrovirus replication. Restriction of retroviral expression in EC cells was studied by using DNA transfection techniques. To investigate the activity of the Moloney murine leukemia virus (M-MuLV)enhancer and promoter sequences, the M-MuLV long terminal repeat and the defined long terminal repeat deletions were linked to neo structural gene sequences that encode resistance to the neomycin analog G418. Transient expression data and drug resistance frequencies support the findings that the M-MuLV enhancer is not active in EC cells but that promoter sequences are functional. In addition, a proviral DNA fragment that encodes the leader RNA sequence of a M-MuLV recombinant retrovirus was found to restrict expression specifically in EC cells. Deletion analysis of the leader fragment localized the inhibitory sequences to a region that spans the M-MuLV tRNA primer binding site. It is not known whether restriction occurs at a transcriptional or posttranscriptional level, but steady-state RNA levels in transient expression assays were significantly reduced.
胚胎癌（EC）细胞对逆转录病毒的复制不敏感。利用DNA转染技术研究了EC细胞中逆转录病毒表达的限制情况。为了研究莫洛尼氏鼠白血病病毒（M-MuLV）增强子和启动子序列的活性，将M-MuLV长末端重复序列和特定的长末端重复序列缺失与编码对新霉素类似物G418抗性的neo结构基因序列相连。瞬时表达数据和耐药频率支持以下发现：M-MuLV增强子在EC细胞中无活性，但启动子序列具有功能。此外，发现一个编码M-MuLV重组逆转录病毒前导RNA序列的前病毒DNA片段在EC细胞中特异性地限制表达。对前导片段的缺失分析将抑制序列定位到跨越M-MuLV tRNA引物结合位点的区域。目前尚不清楚这种限制是发生在转录水平还是转录后水平，但瞬时表达试验中的稳态RNA水平显著降低。
