XRK3F2 Inhibition of p62-ZZ Domain Signaling Rescues Myeloma-Induced GFI1-Driven Epigenetic Repression of the Runx2 Gene in Pre-osteoblasts to Overcome Differentiation Suppression.
XRK3F2 Inhibition of p62-ZZ Domain Signaling Rescues Myeloma-Induced GFI1-Driven Epigenetic Repression of the Runx2 Gene in Pre-osteoblasts to Overcome Differentiation Suppression.
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DOI：
10.3389/fendo.2018.00344
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发表时间：
2018
影响因子：
5.2
通讯作者：
Galson DL
中科院分区：
文献类型：
--
作者：
Adamik J;Silbermann R;Marino S;Sun Q;Anderson JL;Zhou D;Xie XQ;Roodman GD;Galson DL
多发性骨髓瘤骨病（MMBD）的特征是即使在患者达到血液学缓解后仍持续存在的不愈合的溶骨性病变。我们以前报道过骨髓基质细胞（BMSC）中的p62（螯合体-1）是MM诱导的信号复合物介导OB抑制的形成的关键。重要的是，XRK 3F 2，p62-ZZ结构域的抑制剂，在体外减弱MM诱导的Runx 2抑制，并在体内肿瘤存在下诱导新骨形成和重塑。此外，我们报道了MM细胞通过直接结合转录抑制因子GFI 1诱导BMSC中Runx 2基因上抑制性染色质的形成，GFI 1招募组蛋白修饰剂、组蛋白脱乙酰酶1（HDAC 1）和zeste同源物增强子2（EZH 2）。在这项研究中，我们研究了阻断p62-ZZ结构域依赖性信号传导防止MM诱导的BMSC中Runx 2抑制的机制。当存在于MM-preOB共培养物中时，XRK 3F 2阻止MM诱导的Gfi 1上调和Runx 2基因的阻遏。我们还表明，XRK 3F 2阻断p62-ZZ结构域也阻止了MM条件培养基和TNF + IL 7介导的Gfi 1 mRNA上调以及伴随的Runx 2抑制，表明XRK 3F 2对preOB内p62-ZZ结构域信号传导的预防参与了反应。染色质免疫沉淀（ChIP）分析显示，XRK 3F 2降低MM诱导的GFI 1在Runx 2-P1启动子的占有率，并阻止HDAC 1的募集，从而保留了Runx 2上的转录允许染色质标记H3 K9 ac，并允许成骨分化。此外，MM去除后用XRK 3F 2处理MM暴露的preOB减少了Runx 2-P1处的GFI 1富集，并挽救了MM诱导的Runx 2 mRNA及其下游成骨基因靶点的抑制以及成骨分化的增加。此外，与来自健康供体（HD）的hBMSC相比，来自MM患者（MM-hBMSC）的原代BMSC（hBMSC）在成骨条件下几乎没有增加Runx 2启动子上的H3 K9 ac的能力。XRK 3F 2处理使MM-hBMSC中的Runx 2基因H3 K9 ac水平富集至在HD-hBMSC中观察到的水平，但不改变HD-hBMSC H3 K9 ac。重要的是，XRK 3F 2治疗长期MM-hBMSC培养物挽救了成骨分化和矿化。我们的数据表明，用XRK 3F 2阻断p62-ZZ结构域依赖性信号传导可以逆转MM诱导的Runx 2抑制的基于表观遗传的机制，并促进成骨分化。
Multiple myeloma bone disease (MMBD) is characterized by non-healing lytic bone lesions that persist even after a patient has achieved a hematologic remission. We previously reported that p62 (sequestosome-1) in bone marrow stromal cells (BMSC) is critical for the formation of MM-induced signaling complexes that mediate OB suppression. Importantly, XRK3F2, an inhibitor of the p62-ZZ domain, blunted MM-induced Runx2 suppression in vitro, and induced new bone formation and remodeling in the presence of tumor in vivo. Additionally, we reported that MM cells induce the formation of repressive chromatin on the Runx2 gene in BMSC via direct binding of the transcriptional repressor GFI1, which recruits the histone modifiers, histone deacetylase 1 (HDAC1) and Enhancer of zeste homolog 2 (EZH2). In this study we investigated the mechanism by which blocking p62-ZZ domain-dependent signaling prevents MM-induced suppression of Runx2 in BMSC. XRK3F2 prevented MM-induced upregulation of Gfi1 and repression of the Runx2 gene when present in MM-preOB co-cultures. We also show that p62-ZZ-domain blocking by XRK3F2 also prevented MM conditioned media and TNF plus IL7-mediated Gfi1 mRNA upregulation and the concomitant Runx2 repression, indicating that XRK3F2's prevention of p62-ZZ domain signaling within preOB is involved in the response. Chromatin immunoprecipitation (ChIP) analyses revealed that XRK3F2 decreased MM-induced GFI1 occupancy at the Runx2-P1 promoter and prevented recruitment of HDAC1, thus preserving the transcriptionally permissive chromatin mark H3K9ac on Runx2 and allowing osteogenic differentiation. Furthermore, treatment of MM-exposed preOB with XRK3F2 after MM removal decreased GFI1 enrichment at Runx2-P1 and rescued MM-induced suppression of Runx2 mRNA and its downstream osteogenic gene targets together with increased osteogenic differentiation. Further, primary BMSC (hBMSC) from MM patients (MM-hBMSC) had little ability to increase H3K9ac on the Runx2 promoter in osteogenic conditions when compared to hBMSC from healthy donors (HD). XRK3F2 treatment enriched Runx2 gene H3K9ac levels in MM-hBMSC to the level observed in HD-hBMSC, but did not alter HD-hBMSC H3K9ac. Importantly, XRK3F2 treatment of long-term MM-hBMSC cultures rescued osteogenic differentiation and mineralization. Our data show that blocking p62-ZZ domain-dependent signaling with XRK3F2 can reverse epigenetic-based mechanisms of MM-induced Runx2 suppression and promote osteogenic differentiation.