Roulston A, Beauparlant P, Rice N, Hiscott J
Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, Montreal, Quebec, Canada.
J Virol. 1993 Sep;67(9):5235-46. doi: 10.1128/JVI.67.9.5235-5246.1993.
The relationship between human immunodeficiency virus type 1 (HIV-1) infection and the induction of NF-kappa B binding activity was examined in a myeloid cell model of HIV-1 infection derived from the PLB-985 cell line. Chronic infection of PLB-985 cells led to increased monocyte-specific surface marker expression, increased c-fms gene transcription, and morphological alterations consistent with differentiation along the monocytic pathway. PLB-IIIB cells displayed a constitutive NF-kappa B-like binding activity that was distinct from that induced by tumor necrosis factor alpha or phorbol 12-myristate 13-acetate treatment of the parental PLB-985 cell line. This unique DNA binding activity consisted of proteins of 70, 90, and 100 kDa with a high degree of binding specificity for the NF-kappa B site within the PRDII domain of beta interferon. In this report, we characterize the nature of these proteins and demonstrate that binding of these proteins is also induced following Sendai paramyxovirus infection. The 70-kDa protein corresponds to the NF-kappa B RelA (p65) subunit, which is activated in response to an acute paramyxovirus infection or a chronic HIV-1 infection. Virus infection does not appear to alter the amount of RelA (p65) or NFKB1 (p50) but rather affects the capacity of I kappa B alpha to sequester RelA (p65), therefore leading to constitutive levels of RelA DNA binding activity and to increased levels of NF-kappa B-dependent gene activity. The virally induced 90- to 100-kDa proteins have a distinct binding specificity for the PRDII domain and an AT-rich sequence but do not cross-react with NF-kappa B subunit-specific antisera directed against NFKB1 (p105 or p50), NFKB2 (p100 or p52), RelA (p65), or c-rel. DNA binding of the 90- to 100-kDa proteins was not inhibited by recombinant I kappa B alpha/MAD-3 and was resistant to tryptic digestion, suggesting that these proteins may not be NF-kappa B related. Transient cotransfection experiments demonstrated that RelA and NFKB1 expression maximally stimulated HIV-1 LTR- and NF-kappa B-dependent reporter genes; differences in NF-kappa B-like binding activity were also reflected in higher constitutive levels of NF-kappa B-regulated gene expression in HIV-1-infected myeloid cells.
在源自PLB - 985细胞系的HIV - 1感染的髓样细胞模型中，研究了1型人类免疫缺陷病毒（HIV - 1）感染与NF - κB结合活性诱导之间的关系。PLB - 985细胞的慢性感染导致单核细胞特异性表面标志物表达增加、c - fms基因转录增加以及与沿单核细胞途径分化一致的形态学改变。PLB - IIIB细胞表现出一种组成型的NF - κB样结合活性，这与亲本PLB - 985细胞系经肿瘤坏死因子α或佛波酯12 - 肉豆蔻酸酯13 - 乙酸酯处理所诱导的活性不同。这种独特的DNA结合活性由70 kDa、90 kDa和100 kDa的蛋白质组成，对β干扰素PRDII结构域内的NF - κB位点具有高度的结合特异性。在本报告中，我们对这些蛋白质的性质进行了表征，并证明在仙台副粘病毒感染后也会诱导这些蛋白质的结合。70 kDa的蛋白质对应于NF - κB RelA（p65）亚基，它在急性副粘病毒感染或慢性HIV - 1感染时被激活。病毒感染似乎不会改变RelA（p65）或NFKB1（p50）的量，而是影响IκBα隔离RelA（p65）的能力，因此导致RelA DNA结合活性的组成型水平以及NF - κB依赖性基因活性水平的增加。病毒诱导的90至100 kDa蛋白质对PRDII结构域和富含AT的序列具有独特的结合特异性，但不与针对NFKB1（p105或p50）、NFKB2（p100或p52）、RelA（p65）或c - rel的NF - κB亚基特异性抗血清发生交叉反应。90至100 kDa蛋白质的DNA结合不受重组IκBα/MAD - 3抑制，并且对胰蛋白酶消化具有抗性，这表明这些蛋白质可能与NF - κB无关。瞬时共转染实验表明，RelA和NFKB1表达最大程度地刺激了HIV - 1 LTR和NF - κB依赖性报告基因；NF - κB样结合活性的差异也反映在HIV - 1感染的髓样细胞中NF - κB调节基因表达的更高组成型水平上。
