Nuclear Factor of Activated T-cells (NFAT)c2 Inhibits Notch Receptor Signaling in Osteoblasts
Nuclear Factor of Activated T-cells (NFAT)c2 Inhibits Notch Receptor Signaling in Osteoblasts
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DOI：
10.1074/jbc.m112.340455
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发表时间：
2013-01-04
影响因子：
4.8
通讯作者：
Canalis, Ernesto
中科院分区：
文献类型：
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作者：
Zanotti, Stefano;Smerdel-Ramoya, Anna;Canalis, Ernesto
Notch受体调节成骨细胞的发生，Notch激活诱导Notch胞内结构域（NICD）的裂解和核易位，NICD与eb病毒潜伏期c -启动子结合因子-1/无毛/lag-1抑制因子（CSL）相关，并诱导Notch靶基因的转录，如与YRPW基序相关的分裂毛增强子（Hey）1和Hey2。活化t细胞核因子（Nuclear factors of activated t cells， NFAT）是调节破骨细胞发生的转录因子，但其在成骨细胞中的功能尚不清楚。Notch抑制NFATc1转录，但对Notch和NFAT之间的相互作用了解甚少。为了确定Notch对NFAT表达的调控，我们使用了来自Rosa（Notch）小鼠的成骨细胞，其中NICD是在切除loxP侧的STOP磁带后转录的。另外，将野生型C57BL/6成骨细胞暴露于Notch配体delta -样（Dll）1以诱导Notch信号传导或牛血清白蛋白作为对照。在Rosa（Notch）成骨细胞中，Notch通过转录后机制抑制NFATc1的表达，增加Nfatc2 mRNA的表达，而对NFATc3和NFATc4转录没有影响。在暴露于Dll1的C57BL/6成骨细胞中证实了Notch诱导Nfatc2转录本。为了研究NFATc2在成骨细胞中的功能，我们在Rosa（Notch）成骨细胞中过表达构成型活性NFATc2。NFATc2抑制Notch反激活和Hey基因的表达。电泳迁移位移分析显示，NFATc2和CSL结合相似的DNA序列，染色质免疫沉淀表明，NFATc2从Hey2启动子中取代了CSL。NICD和NFATc2对Rosa（Notch）成骨细胞的影响被评估，两种蛋白均抑制成骨细胞功能。总之，Notch稳定Nfatc2转录本，Nfatc2抑制Notch信号传导，两种蛋白均抑制成骨细胞功能。
Notch receptors regulate osteoblastogenesis, and Notch activation induces cleavage and nuclear translocation of the Notch intracellular domain (NICD), which associates with Epstein-Barr virus latency C-promoter binding factor-1/suppressor of hairless/lag-1 (CSL) and induces transcription of Notch target genes, such as hairy enhancer of split-related with YRPW motif (Hey)1 and Hey2. Nuclear factors of activated T-cells (NFAT) are transcription factors that regulate osteoclastogenesis, but their function in osteoblasts is not clear. Notch inhibits NFATc1 transcription, but interactions between Notch and NFAT are understood poorly. To determine the regulation of NFAT expression by Notch, osteoblasts from Rosa(Notch) mice, where NICD is transcribed following excision of a loxP flanked STOP cassette, were used. Alternatively, wild-type C57BL/6 osteoblasts were exposed to the Notch ligand Delta-like (Dll)1 to induce Notch signaling or to bovine serum albumin as control. In Rosa(Notch) osteoblasts, Notch suppressed NFATc1 expression, increased Nfatc2 mRNA by post-transcriptional mechanisms, and had no effect on NFATc3 and NFATc4 transcripts. Induction of Nfatc2 transcripts by Notch was confirmed in C57BL/6 osteoblasts exposed to Dll1. To investigate NFATc2 function in osteoblasts, constitutively active NFATc2 was overexpressed in Rosa(Notch) osteoblasts. NFATc2 suppressed Notch transactivation and expression of Hey genes. Electrophoretic mobility shift assays revealed that NFATc2 and CSL bind to similar DNA sequences, and chromatin immunoprecipitation indicated that NFATc2 displaced CSL from the Hey2 promoter. The effects of NICD and NFATc2 in Rosa(Notch) osteoblasts were assessed, and both proteins inhibited osteoblast function. In conclusion, Notch stabilizes Nfatc2 transcripts, NFATc2 suppresses Notch signaling, and both proteins inhibit osteoblast function.