Adams M, Wong C, Wang D, Romeo J
Center for Biomedical Laboratory Sciences, San Francisco State University, 1600 Holloway Avenue, San Francisco, California 94132, USA.
Virology. 1999 May 10;257(2):397-405. doi: 10.1006/viro.1999.9647.
The ability of HIV to match levels of viral mRNA to the activation state of the host cell may play a role in its ability to persist as well as to replicate. This linkage depends on the function of the viral transcriptional regulatory protein, Tat, which increases the efficiency of RNA elongation (transcriptional processivity) in response to cellular activation. To quantify levels of Tat function in vivo, a quantitative competitive RT-PCR assay was developed that reflects levels of TAR leader fragments (nonprocessive transcripts) and viral mRNA (processive transcripts), indicating low or high levels of Tat function, respectively. The abundance of these RNA species was measured in peripheral blood mononuclear cells (PBMC) of 22 HIV-1-positive individuals (CD4(+) T cell counts 63-934/mm3) and in established cell line models of HIV constitutive replication (H9IIIB) and reversible latency (U1 and ACH-2). In PBMC, the level of total viral transcripts ranged over four orders of magnitude; however, nonprocessive transcription predominated: 70% of PBMC samples had a ratio of processive to total transcripts of <0.3 and none of the samples had 100% processivity. The cell line studies revealed that, even in activated H9IIIB cells, nonprocessive transcription dominates and that latently infected cells can have different transcriptional responses to activation. This is the first study that enumerates degrees of transcriptional processivity in the circulating mononuclear cell compartment and the results suggest that limitation of Tat function may be a common phenotype throughout the course of the disease.
人类免疫缺陷病毒（HIV）使病毒mRNA水平与宿主细胞激活状态相匹配的能力，可能在其持续存在及复制能力方面发挥作用。这种联系取决于病毒转录调节蛋白Tat的功能，Tat可响应细胞激活而提高RNA延伸效率（转录持续性）。为了定量体内Tat功能水平，开发了一种定量竞争性逆转录聚合酶链反应（RT-PCR）检测方法，该方法可反映TAR前导片段（非持续性转录本）和病毒mRNA（持续性转录本）的水平，分别指示Tat功能的低水平或高水平。在22名HIV-1阳性个体（CD4(+)T细胞计数为63 - 934/mm3）的外周血单个核细胞（PBMC）以及HIV组成性复制（H9IIIB）和可逆潜伏（U1和ACH-2）的既定细胞系模型中，测量了这些RNA种类的丰度。在PBMC中，总病毒转录本水平范围跨越四个数量级；然而，非持续性转录占主导：70%的PBMC样本中持续性转录本与总转录本的比率<0.3，且没有样本具有100%的持续性。细胞系研究表明，即使在激活的H9IIIB细胞中，非持续性转录也占主导，并且潜伏感染的细胞对激活可能有不同的转录反应。这是第一项在循环单核细胞区室中列举转录持续性程度的研究，结果表明Tat功能受限可能是疾病全过程中的一种常见表型。
