O'Brien D P, Oltz E M, Van Ness B G
Department of Biochemistry and Institute of Human Genetics, University of Minnesota, Minneapolis 55455, USA.
Mol Cell Biol. 1997 Jul;17(7):3477-87. doi: 10.1128/MCB.17.7.3477.
To further elucidate the potential role of mitogens and cytokines in regulation of the kappa immunoglobulin light-chain locus, we have characterized the activation of transcription factor binding, kappa germ line transcription, DNase I hypersensitivity, and Vkappa-to-Jkappa recombination upon induction of model pre-B-cell lines. We find that both lipopolysaccharide (LPS) and gamma interferon (IFN-gamma) are capable of activating germ line transcription, DNase I hypersensitivity, and recombination of the kappa locus. We also find that transforming growth factor beta is capable of completely inhibiting LPS activation of transcription and recombination but has no apparent effect on activation of transcription factor binding, including activation of NF-kappaB. To address the functional role of NF-kappaB in LPS and IFN-gamma induction of these events, we blocked the nuclear translocation of NF-kappaB by overexpression of a dominant negative mutant of IkappaB-alpha (IkappaB deltaN). Overexpression of the IkappaB deltaN protein results in an inhibition of LPS but not IFN-gamma activation of germ line transcription, DNase I hypersensitivity, and Vkappa-to-Jkappa recombination. Our results demonstrate that activation of NF-kappaB is necessary but not sufficient for LPS activation of transcription and recombination at kappa. These results also suggest that NF-kappaB is not required for IFN-gamma activation of transcription or recombination. These results are important in establishing that there are multiple independent pathways of activation of both transcription and recombination.
为了进一步阐明丝裂原和细胞因子在κ免疫球蛋白轻链基因座调控中的潜在作用，我们对模型前B细胞系诱导后转录因子结合的激活、κ种系转录、DNA酶I超敏反应以及Vκ到Jκ重组进行了表征。我们发现脂多糖（LPS）和γ干扰素（IFN-γ）均能够激活种系转录、DNA酶I超敏反应以及κ基因座的重组。我们还发现转化生长因子β能够完全抑制LPS对转录和重组的激活，但对转录因子结合的激活，包括对NF-κB的激活没有明显影响。为了探讨NF-κB在LPS和IFN-γ诱导这些事件中的功能作用，我们通过过表达IkappaB-α（IkappaB deltaN）的显性负突变体来阻断NF-κB的核转位。IkappaB deltaN蛋白的过表达导致对LPS激活种系转录、DNA酶I超敏反应以及Vκ到Jκ重组的抑制，但对IFN-γ没有抑制作用。我们的结果表明，NF-κB的激活对于LPS激活κ基因座的转录和重组是必要的，但不是充分的。这些结果还表明，NF-κB对于IFN-γ激活转录或重组不是必需的。这些结果对于确定存在多种独立的转录和重组激活途径具有重要意义。
