Tornesello M L, Buonaguro F M, Meglio A, Buonaguro L, Beth-Giraldo E, Giraldo G
Division of Viral Oncology, National Cancer Institute, Fondazione Pascale, Naples, Italy.
J Gen Virol. 1997 Sep;78 ( Pt 9):2199-208. doi: 10.1099/0022-1317-78-9-2199.
Sequence variations in the E6/E7 (nt 34-880) and the L1-(nt 6584-7035) ORFs, and in the long control region (LCR) (nt 7289-93) of human papillomavirus type 16 (HPV-16) were analysed in five penile carcinoma biopsies obtained from Ugandan patients. Uganda is a country with a high incidence of genital cancers. All five isolates were classified as members of African-1 lineage (Af1) by phylogenetic analysis based on LCR sequences. The E6 gene phylogenetic analysis, however, showed that four isolates fell into a new subclass designated Af1-u. This subclass, characterized by three point mutations located at the 5' end of the E6 gene with resulting changes in amino acids at positions 10 and 14, is distinguishable from the Af1 class by the absence of synonymous mutations at nt 286 and 289. The nonsynonymous substitution at nt 335 was present in three out of five samples. The E6 Af1 mutation pattern was present in only a single Ugandan HPV-16 isolate. Nucleotide sequence analysis of the E7 and L1 regions did not allow any Af1 subclass identification. The physical state of the viral DNA in these samples was characterized by PCR and Southern blot analysis. Oligonucleotides which enable amplification of the full length E2 region (nt 2734-3872) failed to amplify the target sequence in four out of five samples, suggesting disruption of the E2 ORF and integration of the HPV genome into the human DNA. Southern blot analysis confirmed the virus integration status. Our results contribute to the characterization of the HPV-16 'African lineages' with the identification of the Af1-u subclass; furthermore, this is also the first report showing that in male genital cancers HPV-16 is integrated into the human genome with disruption of the E2 ORF.
在取自乌干达患者的五份阴茎癌活检样本中，分析了16型人乳头瘤病毒（HPV - 16）的E6/E7（核苷酸34 - 880）和L1（核苷酸6584 - 7035）开放阅读框以及长控制区（LCR，核苷酸7289 - 93）中的序列变异。乌干达是一个生殖器癌症高发的国家。基于LCR序列的系统发育分析显示，所有五个分离株均被归类为非洲-1谱系（Af1）成员。然而，E6基因系统发育分析表明，四个分离株属于一个新的亚类，命名为Af1 - u。该亚类的特征是在E6基因5'端有三个点突变，导致第10和14位氨基酸发生变化，与Af1类的区别在于核苷酸286和289处没有同义突变。五个样本中有三个存在核苷酸335处的非同义替换。E6 Af1突变模式仅在一个乌干达HPV - 16分离株中出现。E7和L1区域的核苷酸序列分析无法识别任何Af1亚类。通过PCR和Southern印迹分析对这些样本中病毒DNA的物理状态进行了表征。能够扩增全长E2区域（核苷酸2734 - 3872）的寡核苷酸在五个样本中有四个未能扩增出目标序列，这表明E2开放阅读框被破坏，HPV基因组整合到了人类DNA中。Southern印迹分析证实了病毒的整合状态。我们的研究结果有助于通过鉴定Af1 - u亚类来表征HPV - 16的“非洲谱系”；此外，这也是第一份表明在男性生殖器癌症中HPV - 16整合到人类基因组且E2开放阅读框被破坏的报告。
