Role of the subcellular localization of ALK tyrosine kinase domain in neuronal differentiation of PC12 cells
Role of the subcellular localization of ALK tyrosine kinase domain in neuronal differentiation of PC12 cells
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DOI：
10.1242/jcs.02695
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发表时间：
2005-12-15
影响因子：
4
通讯作者：
Brunet-de Carvalho, N
中科院分区：
文献类型：
--
作者：
Gouzi, JY;Moog-Lutz, C;Brunet-de Carvalho, N
间变性淋巴瘤激酶（ALK）是一种受体酪氨酸激酶，主要在发育中的中枢和外周神经系统的特定区域瞬时表达。我们先前证明，ALK蛋白酪氨酸激酶（PTK）结构域的膜结合和组成性活性形式通过特异性激活丝裂原活化蛋白激酶（MAP激酶）途径诱导PC 12细胞的神经元样分化。其PTK结构域最初在核-胞质和组成型活性转化蛋白NPM-ALK中鉴定。已经提出参与致癌增殖和存活过程的下游靶标包括磷脂酶C γ（PLC γ）、磷酸肌醇3-激酶（PI 3-激酶）/AKT、STAT 315和Src。因此，我们推测，导致分化或增殖的特定信号通路的激活可以根据ALK PTK结构域的亚细胞定位而受到不同的控制。为了增加其在神经系统中的生理作用的知识，我们集中在目前的研究其亚细胞定位对神经元分化的影响。为了实现这一目标，我们在PC 12细胞中表征了由编码膜结合（通过5811跨膜或肉豆蔻基序列）或胞质ALK衍生蛋白的各种构建体引起的生物学应答和转导途径。为了控制其PTK结构域的激活，我们使用了诱导型二聚化系统。在这里，我们证明了在PC 12细胞中，ALK PTK结构域的膜附着对于通过减少DNA合成来启动神经突生长和增殖抑制至关重要。此外，我们表明，这种分化过程依赖于特异性和持续激活ERK 1/2蛋白。相比之下，该结构域的胞质形式的活化不能诱导MAP激酶活化和细胞分化，但促进PI 3-激酶/AKT依赖性PC 12细胞增殖。这些数据表明，ALK PTK结构域的亚细胞定位是下游转导级联的控制和特异性的决定因素，对于决定神经元细胞的命运至关重要。
Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase essentially and transiently expressed in specific areas of the developing central and peripheral nervous systems. We previously demonstrated that a membrane-bound and constitutively active form of the ALK protein tyrosine kinase (PTK) domain induced the neuron-like differentiation of PC12 cells through specific activation of the mitogen-activated protein kinase (MAP kinase) pathway. Its PTK domain had been originally identified in a nucleo-cytosolic and constitutively active transforming protein, NPM-ALK. Downstream targets involved in oncogenic proliferation and survival processes have been proposed to include phospholipase C gamma (PLC gamma), phosphoinositide 3-kinase (PI 3-kinase)/AKT, STAT 315 and Src. We therefore postulated that activation of specific signaling pathways leading to differentiation or proliferation can be differently controlled depending on the subcellular localization of ALK PTK domain. To increase knowledge of its physiological role in the nervous system, we focused in the present study on the influence of its subcellular localization on neuronal differentiation. To achieve this goal, we characterized biological responses and transduction pathways in PC12 cells elicited by various constructs encoding membrane-bound (through 5811 transmembrane or myristyl sequences) or cytosolic ALK-derived proteins. In order to control the activation of their PTK domain, we used an inducible dimerization system. Here, we demonstrate that membrane attachment of the ALK PTK domain, in PC12 cells, is crucial for initiation of neurite outgrowth and proliferation arrest through a decrease of DNA synthesis. Furthermore, we show that this differentiation process relies on specific and sustained activation of ERK 1/2 proteins. By contrast, activation of the cytosolic form of this domain fails to induce MAP kinase activation and cell differentiation but promotes a PI 3-kinase/AKT-dependant PC12 cell proliferation. These data indicate that subcellular localization of the ALK PTK domain was a determinant for the control and specificity of downstream transduction cascades and was crucial for deciding the fate to which the neuronal cell will be committed.