FAM20C directly binds to and phosphorylates Periostin.
FAM20C directly binds to and phosphorylates Periostin.
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FAM20C直接结合并磷酸化骨膜蛋白。
DOI：
10.1038/s41598-020-74400-6
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发表时间：
2020-10-13
影响因子：
4.6
通讯作者：
Mochida Y
中科院分区：
文献类型：
--
作者：
Lin JH;Lin IP;Ohyama Y;Mochida H;Kudo A;Kaku M;Mochida Y
FAM20 C作为高尔基体酪蛋白激酶发挥功能，并在分泌途径中具有大量激酶底物，这一点已被广泛接受。据报道，FAM20C是维持健康牙周组织所必需的。然而，目前还没有任何报道表明牙周组织中表达的细胞外基质分子确实是FAM 20 C的底物。在这项研究中，我们试图确定FAM20 C的结合伴侣。产生FAM20C野生型（WT）及其激酶无活性形式D478A蛋白。对这些蛋白质进行电泳，并分析考马斯亮蓝（CBB）阳性条带，以通过质谱（MS）分析鉴定FAM20C结合蛋白。通过分析发现了骨膜蛋白，并在细胞培养物和体外研究了FAM20 C和骨膜蛋白之间的结合。我们进一步确定了骨膜蛋白内负责FAM20C结合的结合区域。采用免疫组织化学方法检测小鼠牙周组织中FAM20 C和Periostin的免疫定位。使用骨膜蛋白和FAM20C蛋白进行体外激酶测定以观察FAM20C是否在体外磷酸化骨膜蛋白。我们通过MS分析鉴定了Periostin为FAM20C结合蛋白之一。periostin与FAM 20 C以不依赖于激酶活性的方式相互作用，并且在体外直接结合。我们进一步鉴定了骨膜蛋白中FAM20 C的结合结构域，其定位在骨膜蛋白的成束蛋白（Fas）I结构域1 - 4内。在牙周膜（PDL）细胞外基质中观察到FAM20 C的免疫定位，其中骨膜蛋白在鼠牙周组织中也被免疫染色。FAM20C WT，而不是D478A，在体外磷酸化骨膜蛋白。与FAM20 C和骨膜蛋白的重叠表达模式一致，我们的数据首次证明骨膜蛋白是直接的FAM20 C结合伴侣，并且FAM20 C在体外磷酸化骨膜蛋白。
It is widely accepted that FAM20C functions as a Golgi casein kinase and has large numbers of kinase substrates within the secretory pathway. It has been previously reported that FAM20C is required for maintenance of healthy periodontal tissues. However, there has been no report that any extracellular matrix molecules expressed in periodontal tissues are indeed substrates of FAM20C. In this study, we sought to identify the binding partner(s) of FAM20C. FAM20C wild-type (WT) and its kinase inactive form D478A proteins were generated. These proteins were electrophoresed and the Coomassie Brilliant Blue (CBB)-positive bands were analyzed to identify FAM20C-binding protein(s) by Mass Spectrometry (MS) analysis. Periostin was found by the analysis and the binding between FAM20C and Periostin was investigated in cell cultures and in vitro. We further determined the binding region(s) within Periostin responsible for FAM20C-binding. Immunolocalization of FAM20C and Periostin was examined using mouse periodontium tissues by immunohistochemical analysis. In vitro kinase assay was performed using Periostin and FAM20C proteins to see whether FAM20C phosphorylates Periostin in vitro. We identified Periostin as one of FAM20C-binding proteins by MS analysis. Periostin interacted with FAM20C in a kinase-activity independent manner and the binding was direct in vitro. We further identified the binding domain of FAM20C in Periostin, which was mapped within Fasciclin (Fas) I domain 1–4 of Periostin. Immunolocalization of FAM20C was observed in periodontal ligament (PDL) extracellular matrix where that of Periostin was also immunostained in murine periodontal tissues. FAM20C WT, but not D478A, phosphorylated Periostin in vitro. Consistent with the overlapped expression pattern of FAM20C and Periostin, our data demonstrate for the first time that Periostin is a direct FAM20C-binding partner and that FAM20C phosphorylates Periostin in vitro.
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影响因子：
--
作者：
Acevedo AC;Poulter JA;Alves PG;de Lima CL;Castro LC;Yamaguti PM;Paula LM;Parry DA;Logan CV;Smith CE;Johnson CA;Inglehearn CF;Mighell AJ
通讯作者：
Mighell AJ
影响因子：
7.7
作者：
Cui J;Xiao J;Tagliabracci VS;Wen J;Rahdar M;Dixon JE
通讯作者：
Dixon JE
影响因子：
3.5
作者：
Kantaputra, Piranit Nik;Bongkochwilawan, Chotika;Chaisrisookumporn, Nipon
通讯作者：
Chaisrisookumporn, Nipon
DOI：
10.1016/j.bbrc.2016.01.139
发表时间：
2016-02-19
影响因子：
3.1
作者：
Kii，Isao;Nishiyama，Takashi;Kudo，Akira
通讯作者：
Kudo，Akira
影响因子：
8
作者：
Conway, Simon J.;Izuhara, Kenji;Kudo, Yasusei;Litvin, Judith;Markwald, Roger;Ouyang, Gaoliang;Arron, Joseph R.;Holweg, Cecile T. J.;Kudo, Akira
通讯作者：
Kudo, Akira