Ying B, Smith K, Spindler K R
Department of Genetics, University of Georgia, Athens, Georgia 30602, USA.
J Virol. 1998 Aug;72(8):6325-31. doi: 10.1128/JVI.72.8.6325-6331.1998.
Mouse adenovirus type 1 (MAV-1) mutants with deletions of conserved regions of early region 1A (E1A) or with point mutations that eliminate translation of E1A were used to determine the role of E1A in MAV-1 replication. MAV-1 E1A mutants expressing no E1A protein grew to titers comparable to wild-type MAV-1 titers on mouse fibroblasts (3T6 fibroblasts and fibroblasts derived from Rb+/+, Rb+/-, and Rb-/- transgenic embryos). To test the hypothesis that E1A could induce a quiescent cell to reenter the cell cycle, fibroblasts were serum starved to stop DNA replication and cellular replication and then infected with the E1A mutant and wild-type viruses. All grew to equivalent titers. Steady-state levels of MAV-1 early mRNAs (E1A, E1B, E2, E3, and E4) from 3T6 cells infected with wild-type or E1A mutant virus were examined by Northern analysis. Steady-state levels of mRNAs from the mutant-infected cells were comparable to or greater than the levels found in wild-type virus infections for most of the early regions and for two late genes. The E2 mRNA levels were slightly reduced in all mutant infections relative to wild-type infections. E1A mRNA was not detected from infections with the MAV-1 E1A null mutant, pmE109, or from infections with similar MAV-1 E1A null mutants, pmE112 and pmE113. The implications for the lack of a requirement of E1A in cell culture are discussed.
1型小鼠腺病毒（MAV-1）早期区域1A（E1A）保守区域缺失的突变体或消除E1A翻译的点突变体被用于确定E1A在MAV-1复制中的作用。不表达E1A蛋白的MAV-1 E1A突变体在小鼠成纤维细胞（3T6成纤维细胞以及源自Rb+/+、Rb+/−和Rb−/−转基因胚胎的成纤维细胞）上生长至与野生型MAV-1滴度相当的滴度。为了验证E1A可诱导静止细胞重新进入细胞周期这一假说，将成纤维细胞进行血清饥饿处理以停止DNA复制和细胞增殖，然后用E1A突变体和野生型病毒进行感染。所有病毒均生长至等效滴度。通过Northern分析检测了感染野生型或E1A突变体病毒的3T6细胞中MAV-1早期mRNA（E1A、E1B、E2、E3和E4）的稳态水平。对于大多数早期区域和两个晚期基因，突变体感染细胞的mRNA稳态水平与野生型病毒感染中的水平相当或更高。相对于野生型感染，所有突变体感染中E2 mRNA水平均略有降低。在MAV-1 E1A缺失突变体pmE109感染中或在类似的MAV-1 E1A缺失突变体pmE112和pmE113感染中均未检测到E1A mRNA。文中讨论了在细胞培养中不需要E1A的意义。
