Buchholz Ursula J, Biacchesi Stéphane, Pham Quynh N, Tran Kim C, Yang Lijuan, Luongo Cindy L, Skiadopoulos Mario H, Murphy Brian R, Collins Peter L
Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892-8007, USA.
J Virol. 2005 Jun;79(11):6588-97. doi: 10.1128/JVI.79.11.6588-6597.2005.
The M2 gene of human metapneumovirus (HMPV) contains two overlapping open reading frames (ORFs), M2-1 and M2-2. The expression of separate M2-1 and M2-2 proteins from these ORFs was confirmed, and recombinant HMPVs were recovered in which expression of M2-1 and M2-2 was ablated individually or together [rdeltaM2-1, rdeltaM2-2, and rdeltaM2(1+2)]. Each M2 mutant virus directed efficient multicycle growth in Vero cells. The ability to recover HMPV lacking M2-1 contrasts with human respiratory syncytial virus, for which M2-1 is an essential transcription factor. Expression of the downstream HMPV M2-2 ORF was not reduced when translation of the upstream M2-1 ORF was silenced, indicating that it is initiated separately. The rdeltaM2-2 mutants exhibited a two- to fivefold increase in the accumulation of mRNA, normalized to the genome template, suggesting that M2-2 has a role in regulating RNA synthesis. Replication and immunogenicity were tested in hamsters. Animals infected intranasally with rdeltaM2-1 or rdeltaM2(1+2) did not have recoverable virus in the lungs or nasal turbinates on days 3 or 5 postinfection and did not develop HMPV-neutralizing serum antibodies or resistance to HMPV challenge. Thus, M2-1 appears to be essential for significant virus replication in vivo. In animals infected with rdeltaM2-2, virus was recovered from only 1 of 12 animals and only in the nasal turbinates on a single day. However, all of the animals developed a high titer of HMPV-neutralizing serum antibodies and were highly protected against challenge with wild-type HMPV. The HMPV rdeltaM2-2 virus is a promising and highly attenuated HMPV vaccine candidate.
人偏肺病毒（HMPV）的M2基因包含两个重叠的开放阅读框（ORF），即M2-1和M2-2。已证实从这些ORF可分别表达M2-1和M2-2蛋白，并获得了重组HMPV，其中M2-1和M2-2的表达分别或共同被消除[rdeltaM2-1、rdeltaM2-2和rdeltaM2(1+2)]。每种M2突变病毒在Vero细胞中均能高效进行多轮生长。恢复缺乏M2-1的HMPV的能力与人类呼吸道合胞病毒形成对比，后者的M2-1是一种必需的转录因子。当上游M2-1 ORF的翻译被沉默时，下游HMPV M2-2 ORF的表达并未降低，这表明它是单独起始的。rdeltaM2-2突变体在以基因组模板标准化后的mRNA积累量增加了2至5倍，这表明M2-2在调节RNA合成中发挥作用。在仓鼠中测试了复制和免疫原性。经鼻感染rdeltaM2-1或rdeltaM2(1+2)的动物在感染后第3天或第5天，肺或鼻甲中未检测到可恢复的病毒，也未产生HMPV中和血清抗体或对HMPV攻击的抵抗力。因此，M2-1似乎对于体内显著的病毒复制至关重要。在感染rdeltaM2-2的动物中，仅从12只动物中的1只身上回收了病毒，且仅在某一天从鼻甲中检测到。然而，所有动物均产生了高滴度的HMPV中和血清抗体，并对野生型HMPV攻击具有高度抵抗力。HMPV rdeltaM2-2病毒是一种有前景的、高度减毒的HMPV疫苗候选株。
