Osteoblast regulation via ligand-activated nuclear trafficking of the oxytocin receptor
Osteoblast regulation via ligand-activated nuclear trafficking of the oxytocin receptor
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DOI：
10.1073/pnas.1419349111
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发表时间：
2014-11-18
影响因子：
11.1
通讯作者：
Zaidi, Mone
中科院分区：
文献类型：
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作者：
Di Benedetto, Adriana;Sun, Li;Zaidi, Mone
我们报告，催产素（Oxt）受体（Oxtrs），刺激的配体Oxt，易位到成骨细胞的细胞核，暗示这一过程中的行动，Oxt成骨细胞的成熟。连续的完整细胞或分离的核质体剥离的外核膜的免疫细胞化学显示渐进的OXTR核定位，这种核转位证实了通过监测的运动的OXTR-绿色荧光蛋白，以及通过免疫金标记。核Oxtr定位通过核裂解物蛋白质的Western免疫印迹和MS最终显示。我们发现，Oxtrs进入细胞核的通道是通过与β-抑制蛋白（Arrbs），小GTTRB Rab 5，输入蛋白-β（Kpnb 1）和转运蛋白-1（Tnpo 1）的连续相互作用来促进的。siRNA介导的Arrb 1、Arrb 2或Tnpo 1的敲除废除了Oxt诱导的成骨细胞分化基因osterix（Sp 7）、Atf 4、骨唾液蛋白（Ibsp）和骨钙素（Bglap）的表达，而不影响Erk磷酸化。同样地，在不影响pErk的情况下，通过将核定位信号的富含Ser的簇突变为Ala来抑制Arrb募集，从而废除了核输入和Oxtr诱导的基因表达。这些研究定义了一个以前未确定的机制OXTR作用于骨和开放的可能性直接转录调节核G蛋白偶联受体。
We report that oxytocin (Oxt) receptors (Oxtrs), on stimulation by the ligand Oxt, translocate into the nucleus of osteoblasts, implicating this process in the action of Oxt on osteoblast maturation. Sequential immunocytochemistry of intact cells or isolated nucleoplasts stripped of the outer nuclear membrane showed progressive nuclear localization of the Oxtr; this nuclear translocation was confirmed by monitoring the movement of Oxtr-EGFP as well as by immunogold labeling. Nuclear Oxtr localization was conclusively shown by Western immunoblotting and MS of nuclear lysate proteins. We found that the passage of Oxtrs into the nucleus was facilitated by successive interactions with beta-arrestins (Arrbs), the small GTPase Rab5, importin-beta (Kpnb1), and transportin-1 (Tnpo1). siRNA-mediated knockdown of Arrb1, Arrb2, or Tnpo1 abrogated Oxt-induced expression of the osteoblast differentiation genes osterix (Sp7), Atf4, bone sialoprotein (Ibsp), and osteocalcin (Bglap) without affecting Erk phosphorylation. Likewise and again, without affecting pErk, inhibiting Arrb recruitment by mutating Ser rich clusters of the nuclear localization signal to Ala abolished nuclear import and Oxtr-induced gene expression. These studies define a previously unidentified mechanism for Oxtr action on bone and open possibilities for direct transcriptional modulation by nuclear G protein-coupled receptors.