Chalifour L E, Wirak D O, Wassarman P M, DePamphilis M L
J Virol. 1986 Sep;59(3):619-27. doi: 10.1128/JVI.59.3.619-627.1986.
Simian virus 40 (SV40) large- and small-tumor antigens (T-Ag, t-Ag) are normally synthesized early after infection of either permissive (monkey) or nonpermissive (mouse) fibroblasts, whereas an equivalent amount of viral coat protein (V-Ag) is observed late after infection of permissive cells and only after viral DNA replication has occurred. To determine whether or not expression of these genes is regulated in the same manner during early mammalian development, SV40 DNA was injected into the nuclei of mouse oocytes and one- and two-cell embryos. In oocytes, about three times more V-Ag was produced than T-Ag, and both were synthesized concomitantly in the same cells. Viral mRNA and proteins synthesized in oocytes comigrated during gel electrophoresis with the same products synthesized in SV40-infected monkey cells. Viral gene expression required circular DNA molecules injected into the nuclei of transcriptionally and translationally active cells. Injected DNA was stable and underwent conformational changes consistent with chromatin assembly. Oocytes did not replicate either polyomavirus or SV40 DNA. Thus, the temporal order of viral gene expression is circumvented in mouse germ cells, allowing these proteins to be expressed concurrently and in equivalent amounts with no requirement for DNA replication. However, in preimplantation embryos, neither T-Ag nor V-Ag was detected by immunoprecipitation although T-Ag synthesis was demonstrated as a specific requirement for SV40 DNA replication. Thus, viral gene expression in mouse embryos as early as the one-cell stage was reduced at least 500-fold relative to that in oocytes. Similarities between SV40 gene expression in mouse oocytes and that in Xenopus oocytes suggest that germ cells in higher animals share common regulatory mechanisms.
猴病毒40(SV40)大肿瘤抗原和小肿瘤抗原(T抗原、t抗原)通常在允许性(猴)或非允许性(小鼠)成纤维细胞感染后早期合成,而在允许性细胞感染后期且仅在病毒DNA复制发生后才观察到等量的病毒衣壳蛋白(V抗原)。为了确定这些基因在早期哺乳动物发育过程中是否以相同方式受到调控,将SV40 DNA注射到小鼠卵母细胞以及单细胞和双细胞胚胎的细胞核中。在卵母细胞中,产生的V抗原比T抗原多约三倍,并且两者在同一细胞中同时合成。卵母细胞中合成的病毒mRNA和蛋白质在凝胶电泳中与SV40感染的猴细胞中合成的相同产物迁移情况一致。病毒基因表达需要将环状DNA分子注射到具有转录和翻译活性的细胞核中。注射的DNA是稳定的,并经历了与染色质组装一致的构象变化。卵母细胞不复制多瘤病毒或SV40 DNA。因此,在小鼠生殖细胞中规避了病毒基因表达的时间顺序,使得这些蛋白质能够同时且等量表达,而无需DNA复制。然而,在植入前胚胎中,通过免疫沉淀未检测到T抗原或V抗原,尽管T抗原的合成被证明是SV40 DNA复制的特定要求。因此,早在单细胞阶段的小鼠胚胎中的病毒基因表达相对于卵母细胞至少降低了500倍。小鼠卵母细胞和非洲爪蟾卵母细胞中SV40基因表达的相似性表明高等动物的生殖细胞具有共同的调控机制。