Lewinski M K, Bisgrove D, Shinn P, Chen H, Hoffmann C, Hannenhalli S, Verdin E, Berry C C, Ecker J R, Bushman F D
Infectious Disease Laboratory, The Salk Institute, La Jolla, California 92037, USA.
J Virol. 2005 Jun;79(11):6610-9. doi: 10.1128/JVI.79.11.6610-6619.2005.
We have investigated regulatory sequences in noncoding human DNA that are associated with repression of an integrated human immunodeficiency virus type 1 (HIV-1) promoter. HIV-1 integration results in the formation of precise and homogeneous junctions between viral and host DNA, but integration takes place at many locations. Thus, the variation in HIV-1 gene expression at different integration sites reports the activity of regulatory sequences at nearby chromosomal positions. Negative regulation of HIV transcription is of particular interest because of its association with maintaining HIV in a latent state in cells from infected patients. To identify chromosomal regulators of HIV transcription, we infected Jurkat T cells with an HIV-based vector transducing green fluorescent protein (GFP) and separated cells into populations containing well-expressed (GFP-positive) or poorly expressed (GFP-negative) proviruses. We then determined the chromosomal locations of the two classes by sequencing 971 junctions between viral and cellular DNA. Possible effects of endogenous cellular transcription were characterized by transcriptional profiling. Low-level GFP expression correlated with integration in (i) gene deserts, (ii) centromeric heterochromatin, and (iii) very highly expressed cellular genes. These data provide a genome-wide picture of chromosomal features that repress transcription and suggest models for transcriptional latency in cells from HIV-infected patients.
我们研究了人类非编码DNA中的调控序列，这些序列与整合的1型人类免疫缺陷病毒（HIV-1）启动子的抑制有关。HIV-1整合导致病毒与宿主DNA之间形成精确且均匀的连接，但整合发生在许多位置。因此，HIV-1基因在不同整合位点的表达差异反映了附近染色体位置调控序列的活性。由于HIV转录的负调控与在感染患者细胞中使HIV维持潜伏状态有关，因此其特别受关注。为了鉴定HIV转录的染色体调节因子，我们用转导绿色荧光蛋白（GFP）的基于HIV的载体感染Jurkat T细胞，并将细胞分为含有高表达（GFP阳性）或低表达（GFP阴性）前病毒的群体。然后，我们通过对病毒与细胞DNA之间的971个连接进行测序来确定这两类细胞的染色体位置。通过转录谱分析来表征内源性细胞转录的可能影响。低水平的GFP表达与整合到（i）基因荒漠、（ii）着丝粒异染色质和（iii）高表达的细胞基因中相关。这些数据提供了全基因组范围内抑制转录的染色体特征图谱，并为HIV感染患者细胞中的转录潜伏期提出了模型。
