Bishop Kathleen A, Wang Xiaohua, Coy Heidi M, Meyer Mark B, Gumperz Jenny E, Pike J Wesley
Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706.
J Cell Biochem. 2015 Feb;116(2):320-30. doi: 10.1002/jcb.24974.
In addition to osteoblast lineage cells, the TNF-like factor receptor activator of NF-κB ligand (RANKL) is expressed in both B and T cells and may play a role in bone resorption. Rankl gene (Tnfsf11) expression in mouse T cells is mediated through multiple distal elements marked by increased transcription factor occupancy, histone tail acetylation, and RNA polymerase II recruitment. Little is known, however, of the regulation of human TNFSF11 in T cells. Accordingly, we examined the consequence of T cell activation on the expression of this factor both in Jurkat cells and in primary human T cells. We then explored the mechanism of this regulation by scanning over 400 kb of DNA surrounding the TNFSF11 locus for regulatory enhancers using ChIP-chip analysis. Histone H3/H4 acetylation enrichment identified putative regulatory regions located between -170 and -220 kb upstream of the human TNFSF11 TSS that we designated the human T cell control region (hTCCR). This region showed high sequence conservation with the mouse TCCR. Inhibition of MEK1/2 by U0126 resulted in decreased RANKL expression suggesting that stimulation through MEK1/2 was a prerequisite. ChIP-chip analysis also revealed that c-FOS was recruited to the hTCCR as well. Importantly, both the human TNFSF11 D5a/b (RLD5a/b) enhancer and segments of the hTCCR mediated robust inducible reporter activity following TCR activation. Finally, SNPs implicated in diseases characterized by dysregulated BMD co-localized to the hTCCR region. We conclude that the hTCCR region contains a cell-selective set of enhancers that plays an integral role in the transcriptional regulation of the TNFSF11 gene in human T cells.
除了成骨细胞系细胞外，核因子κB受体活化因子配体（RANKL）这种肿瘤坏死因子样因子受体也在B细胞和T细胞中表达，并且可能在骨吸收中发挥作用。小鼠T细胞中的Rankl基因（Tnfsf11）表达是通过多个远端元件介导的，这些元件的特征是转录因子占据增加、组蛋白尾部乙酰化以及RNA聚合酶II募集。然而，关于人类TNFSF11在T细胞中的调控知之甚少。因此，我们研究了T细胞活化对Jurkat细胞和原代人T细胞中该因子表达产生的影响。然后，我们通过染色质免疫沉淀芯片分析（ChIP-chip分析）扫描TNFSF11基因座周围超过400kb的DNA区域以寻找调控增强子，从而探究这种调控的机制。组蛋白H3/H4乙酰化富集鉴定出位于人类TNFSF11转录起始位点（TSS）上游-170至-220kb之间的假定调控区域，我们将其命名为人类T细胞控制区域（hTCCR）。该区域与小鼠TCCR具有高度的序列保守性。U0126对MEK1/2的抑制导致RANKL表达降低，这表明通过MEK1/2进行刺激是一个先决条件。ChIP-chip分析还显示c-FOS也被募集到hTCCR。重要的是，人类TNFSF11 D5a/b（RLD5a/b）增强子和hTCCR的片段在TCR激活后介导了强大的诱导型报告基因活性。最后，与骨密度失调相关疾病有关的单核苷酸多态性（SNP）共定位于hTCCR区域。我们得出结论，hTCCR区域包含一组细胞选择性增强子，它们在人类T细胞中TNFSF11基因的转录调控中发挥着不可或缺的作用。
