Chakraborti Samitabh, Banerjea Akhil C
Laboratory of Virology, National Institute of Immunology, JNU Campus, Aruna Asaf Ali Marg, 110067, New Delhi, India.
Mol Ther. 2003 Jun;7(6):817-26. doi: 10.1016/s1525-0016(03)00096-0.
Nucleic acid-based antiviral approaches have been tried against multiple HIV-1 genes with the purpose of down-regulating its replication. A unique stem-loop structure called TAR is present at the 5'-end of all HIV-1 transcripts; Tat and other cellular proteins bind to TAR and thus govern transcription. Therefore, HIV-1 TAR is an attractive target against which various antiviral approaches could be tried. We screened several DNA enzymes (Dz's) containing the 10-23 catalytic motif and a single Dz containing the 8-17 catalytic motif against the HIV-1 TAR RNA. Dz's directed against the predicted single-stranded bulge regions showed sequence-specific cleavage activities. One of the two Dz's, namely Dz-475, showed moderate cleavage activity in complete absence of Mg(2+). Addition of unrelated sequences at the 5'-end of the HIV-1 TAR RNA rendered it susceptible to four additional Dz-mediated cleavages. Both Dz's (470 and 475) showed significant intracellular reduction of HIV-1 gene expression. Dz-475-treated cells showed significant protection against T-tropic and macrophage-tropic HIV-1 challenge. Dz-475-transfected T-lymphocytes, human PBMCs, or chronically infected cell lines showed marked viral resistance. Unique features of this antiviral strategy with respect to HIV-1 gene inhibition are discussed.
基于核酸的抗病毒方法已针对多种HIV-1基因进行尝试，目的是下调其复制。在所有HIV-1转录本的5'端存在一种称为TAR的独特茎环结构；Tat和其他细胞蛋白与TAR结合，从而控制转录。因此，HIV-1 TAR是一个有吸引力的靶点，可以尝试各种抗病毒方法。我们针对HIV-1 TAR RNA筛选了几种含有10-23催化基序的DNA酶（Dz）和一种含有8-17催化基序的单一Dz。针对预测的单链凸起区域的Dz显示出序列特异性切割活性。两种Dz中的一种，即Dz-475，在完全不存在Mg(2+)的情况下显示出适度的切割活性。在HIV-1 TAR RNA的5'端添加无关序列使其易受另外四种Dz介导的切割。两种Dz（470和475）均显示出HIV-1基因表达在细胞内显著降低。经Dz-475处理的细胞对T嗜性和巨噬细胞嗜性HIV-1攻击显示出显著的保护作用。转染了Dz-475的T淋巴细胞、人外周血单核细胞或慢性感染细胞系显示出明显的病毒抗性。讨论了这种针对HIV-1基因抑制的抗病毒策略的独特特征。
