Phosphorylation of a splice variant of collapsin response mediator protein 2 in the nucleus of tumour cells links cyclin dependent kinase-5 to oncogenesis.
Phosphorylation of a splice variant of collapsin response mediator protein 2 in the nucleus of tumour cells links cyclin dependent kinase-5 to oncogenesis.
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DOI：
10.1186/s12885-015-1691-1
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发表时间：
2015-11-10
期刊：
影响因子：
3.8
通讯作者：
Sutherland C
中科院分区：
文献类型：
--
作者：
Grant NJ;Coates PJ;Woods YL;Bray SE;Morrice NA;Hastie CJ;Lamont DJ;Carey FA;Sutherland C
细胞周期蛋白依赖性蛋白激酶-5（CDK 5）是CDK家族的一个不寻常的成员，因为它不受细胞周期调节。然而，它的许多底物在细胞生长和肿瘤发生中起作用，提高了CDK 5调节可能具有治疗益处的可能性。为了确定CDK 5活性的变化是否与肿瘤发生相关，可以与适当的对照相比定量疾病组织中CDK 5靶标的磷酸化。然而，生理学和病理生理学CDK 5底物的身份仍然是争论的主题，使得CDK 5活性生物标志物的选择变得困难。在这里，我们使用在体外和细胞磷酸化测定，以确定新的功能，赋予增强CDK 5的选择性CDK 5的靶序列决定簇，提供手段，选择底物生物标志物的CDK 5活性更有信心。然后，我们表征了我们确定的最佳CDK 5底物的工具，以监测其在人体组织中的磷酸化，并使用这些来询问人类肿瘤阵列。Arg/Lys氨基酸和脯氨酸的N-末端两个残基与磷酸化残基的紧密接近都改善了CDK 5对底物的识别。相比之下，靶残基C-末端的脯氨酸两个残基的存在显著降低磷酸化速率。Collapsin Response Mediator-2（CRMP 2）的Serine-522是一种经过验证的CDK 5底物，具有许多这些结构标准。我们生成并鉴定了Ser 522的磷酸化特异性抗体，并表明磷酸化出现在人类肿瘤（肺、乳腺和淋巴瘤）中，与周围的非肿瘤组织形成鲜明对比。在肺癌中，抗磷酸化Ser 522信号在鳞状细胞癌中比腺癌更常见。最后，我们证明，这是一个特定的和不寻常的剪接变体的CRMP 2（CRMP 2A），是在肿瘤细胞中磷酸化。这一数据首次将CDK 5底物磷酸化改变与某些但非所有肿瘤类型的肿瘤发生联系起来，表明CDK 5活性改变与发病机制有关。这些数据确定了一种新的致癌机制，其中CDK 5激活诱导肿瘤细胞核中的CRMP 2A磷酸化。本文的在线版本（doi：10.1186/s12885-015-1691-1）包含补充材料，可供授权用户使用。
Cyclin-dependent protein kinase-5 (CDK5) is an unusual member of the CDK family as it is not cell cycle regulated. However many of its substrates have roles in cell growth and oncogenesis, raising the possibility that CDK5 modulation could have therapeutic benefit. In order to establish whether changes in CDK5 activity are associated with oncogenesis one could quantify phosphorylation of CDK5 targets in disease tissue in comparison to appropriate controls. However the identity of physiological and pathophysiological CDK5 substrates remains the subject of debate, making the choice of CDK5 activity biomarkers difficult. Here we use in vitro and in cell phosphorylation assays to identify novel features of CDK5 target sequence determinants that confer enhanced CDK5 selectivity, providing means to select substrate biomarkers of CDK5 activity with more confidence. We then characterize tools for the best CDK5 substrate we identified to monitor its phosphorylation in human tissue and use these to interrogate human tumour arrays. The close proximity of Arg/Lys amino acids and a proline two residues N-terminal to the phosphorylated residue both improve recognition of the substrate by CDK5. In contrast the presence of a proline two residues C-terminal to the target residue dramatically reduces phosphorylation rate. Serine-522 of Collapsin Response Mediator-2 (CRMP2) is a validated CDK5 substrate with many of these structural criteria. We generate and characterise phosphospecific antibodies to Ser522 and show that phosphorylation appears in human tumours (lung, breast, and lymphoma) in stark contrast to surrounding non-neoplastic tissue. In lung cancer the anti-phospho-Ser522 signal is positive in squamous cell carcinoma more frequently than adenocarcinoma. Finally we demonstrate that it is a specific and unusual splice variant of CRMP2 (CRMP2A) that is phosphorylated in tumour cells. For the first time this data associates altered CDK5 substrate phosphorylation with oncogenesis in some but not all tumour types, implicating altered CDK5 activity in aspects of pathogenesis. These data identify a novel oncogenic mechanism where CDK5 activation induces CRMP2A phosphorylation in the nuclei of tumour cells. The online version of this article (doi:10.1186/s12885-015-1691-1) contains supplementary material, which is available to authorized users.