Adelson M E, Martinand-Mari C, Iacono K T, Muto N F, Suhadolnik R J
Fels Institute for Cancer Research, Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA, USA.
Eur J Biochem. 1999 Sep;264(3):806-15. doi: 10.1046/j.1432-1327.1999.00661.x.
Current strategies against the human immunodeficiency virus type 1 (HIV-1), including nucleoside analogues and protease inhibitors, have limited effectiveness as shown by the evolution of resistant retroviral strains and the presence of latent HIV-1 reservoirs. Therefore, it is necessary to look beyond anti-retroviral strategies and to rely on the body's immune system to inhibit HIV-1 replication. In this study, we approach the inhibition of HIV-1 replication by upregulation of the antiviral pathway that is natural to mammalian cells. Vectors were constructed which were capable of transferring the antiviral enzyme, p68 kinase (PKR), into target SupT1 lymphoblastoid cells under HIV-1 LTR transcriptional regulation via a retroviral-mediated shuttle system. We report a significant inhibition of HIV-1 replication in HIV-1 LTR-PKR cDNA transduced clones (105-10 : 239 and 106-4 : 560) expressing different PKR levels as measured by inhibition of HIV-1 induced syncytia formation and HIV-1 reverse transcriptase activity. Whereas the expression of PKR in parental vector transduced clone (N2-20P) is down-regulated 48 h after HIV-1 infection, the two transduced clones (one with PKR in the forward orientation and one in the reverse orientation) demonstrate increased PKR expression through 96 h post-infection, concomitant with an increase in eIF-2alpha phosphorylation and an increase in NF-kappaB activity at 72 h postinfection. These results demonstrate that the overexpression of PKR can inhibit HIV-1 replication and confirm the involvement of PKR in the IFN-associated antiviral pathway against HIV-1 infection. Finally, the treatment of the transduced clone 106-4 : 560 with AZT resulted in complete inhibition of HIV-1 replication.
目前针对1型人类免疫缺陷病毒（HIV-1）的策略，包括核苷类似物和蛋白酶抑制剂，其有效性有限，耐药逆转录病毒株的出现以及潜伏HIV-1储存库的存在就表明了这一点。因此，有必要超越抗逆转录病毒策略，依靠人体免疫系统来抑制HIV-1复制。在本研究中，我们通过上调哺乳动物细胞天然存在的抗病毒途径来抑制HIV-1复制。构建了载体，其能够通过逆转录病毒介导的穿梭系统在HIV-1长末端重复序列（LTR）转录调控下将抗病毒酶p68激酶（PKR）转移到靶标SupT1淋巴母细胞中。我们报告称，在通过抑制HIV-1诱导的合胞体形成和HIV-1逆转录酶活性来测量的情况下，表达不同PKR水平的HIV-1 LTR-PKR cDNA转导克隆（105-10 : 239和106-4 : 560）中HIV-1复制受到显著抑制。亲本载体转导克隆（N2-20P）中PKR的表达在HIV-1感染后48小时下调，而两个转导克隆（一个PKR为正向，一个为反向）在感染后96小时内PKR表达增加，同时在感染后72小时eIF-2α磷酸化增加且NF-κB活性增加。这些结果表明PKR的过表达可以抑制HIV-1复制，并证实PKR参与了针对HIV-1感染的干扰素相关抗病毒途径。最后，用齐多夫定（AZT）处理转导克隆106-4 : 560导致HIV-1复制完全受到抑制。
