Rossi C, Gibellini D, Barbanti-Brodano G, Betti M, Boarini C, Pengue G, Lania L, Caputo A
Department of Experimental and Diagnostic Medicine, University of Ferrara, Italy.
J Med Virol. 1999 Jul;58(3):264-72. doi: 10.1002/(sici)1096-9071(199907)58:3<264::aid-jmv12>3.0.co;2-v.
It has been demonstrated previously that the transcriptional repressor domain called the Krüppel-associated box (KRAB), conserved in a large number of Krüppel-type zinc finger proteins, fused to Tat transdominant negative mutants, is able to silence HIV-1 long terminal repeat (LTR)-driven gene expression in transient transfection assays. In the present study chimeric Tat mutant-KRAB retroviral expression vectors were used to control HIV-1 replication in acutely infected cells. It was found that while transient and stable expression of Tat mutant-KRAB chimeric proteins represses HIV-1 LTR-driven gene transcription in transient assays, stable expression of Tat mutant-KRAB chimeric molecules does not confer resistance to HIV-1 infection in Jurkat T lymphocytic cell lines. The results provide further evidence that transient transfection may underestimate the role of chromosomal structure in transcriptional regulation and highlight the caveat of direct extrapolation of transient results for designing gene therapy strategies for efficient control of HIV-1 infection.
先前已经证明，在大量克氏锥虫型锌指蛋白中保守的称为克氏锥虫相关盒（KRAB）的转录抑制结构域，与Tat反式显性负突变体融合后，能够在瞬时转染试验中沉默HIV-1长末端重复序列（LTR）驱动的基因表达。在本研究中，嵌合Tat突变体-KRAB逆转录病毒表达载体用于控制急性感染细胞中的HIV-1复制。结果发现，虽然Tat突变体-KRAB嵌合蛋白的瞬时和稳定表达在瞬时试验中抑制HIV-1 LTR驱动的基因转录，但Tat突变体-KRAB嵌合分子的稳定表达并不能使Jurkat T淋巴细胞系对HIV-1感染产生抗性。这些结果进一步证明，瞬时转染可能低估了染色体结构在转录调控中的作用，并突出了直接外推瞬时结果以设计有效控制HIV-1感染的基因治疗策略的警告。
