Mobility of TrkA is regulated by phosphorylation and interactions with the low-affinity NGF receptor.
Mobility of TrkA is regulated by phosphorylation and interactions with the low-affinity NGF receptor.
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TrkA 的移动性通过磷酸化以及与低亲和力 NGF 受体的相互作用来调节。
DOI：
10.1021/bi9719253
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Ross,AH
中科院分区：
文献类型：
--
作者：
Wolf,DE;McKinnon-Thompson,C;Daou,MC;Stephens,RM;Kaplan,DR;Ross,AH
神经生长因子（NGF）受体是两种蛋白质gp 75和酪氨酸激酶TrkA的复合物。利用光漂白后的荧光恢复，我们研究了TrkA受体的扩散特性。在同时表达gp 75和TrkA的PC 12细胞中，TrkA相对固定，只有28 ± 1%的受体分子自由扩散，D =（3.64 ± 0.23）× 10- 9 cm 2/s。加入神经生长因子后，细胞移动的比例降至21 ± 1%，D =（4.11 ± 0.18）× 10- 9 cm 2/s。使用Sf 9杆状病毒表达系统，我们能够在gp 75存在和不存在的情况下研究TrkA。在Sf 9细胞上，TrkA在无gp 75存在时的移动的率为46 ± 2%，D =（2.64 ± 0.21）× 10 ~（-9）cm ~ 2/s;在有gp 75存在时的移动率为43 ± 2%，D =（2.31 ± 0.25）× 10 ~（-9）cm ~ 2/s。因此，gp 75不改变TrkA迁移率。在gp 75不存在和存在的情况下，向培养基中加入NGF使TrkA的移动的部分大约减半。然而，使用TrkA的激酶缺陷突变体，我们发现配体诱导的固定化需要在gp 75的情况下而不是在其存在下的活性激酶。此外，使用点突变在特定的TrkA自磷酸化位点，我们确定，流动性是由多个磷酸化位点控制，但在Y 490的SHC结合位点可能是特别重要的配体诱导的固定化的TrkA。因此，两种机制导致NGF诱导的TrkA的固定化，第一种是由TrkA的自磷酸化引起的，第二种是通过TrkA与gp 75的结合而发生的。
The nerve growth factor (NGF) receptor is a complex of two proteins, gp75 and the tyrosine kinase TrkA. Using fluorescence recovery after photobleaching, we have studied the diffusion properties of the TrkA receptor. For PC12 cells that express both gp75 and TrkA, TrkA was relatively immobile with only 28 ± 1% of receptor molecules free to diffuse withD= (3.64 ± 0.23) × 10-9cm2/s. Addition of NGF decreased the mobile fraction to 21 ± 1% withD= (4.11 ± 0.18) × 10-9cm2/s. Using the Sf9 baculovirus expression system, we were able to study TrkA in the absence and presence of gp75. On Sf9 cells, TrkA showed a mobile fraction of 46 ± 2% withD= (2.64 ± 0.21) × 10-9cm2/s in the absence of gp75 and 43 ± 2% withD= (2.31 ± 0.25) × 10-9cm2/s in its presence. Thus, gp75 did not alter TrkA mobility. Addition of NGF to the medium approximately halved the mobile fraction for TrkA in both the absence and presence of gp75. However, using a kinase-deficient mutant of TrkA, we found that ligand-induced immobilization requires an active kinase in the absence of gp75 but not in its presence. In addition, using point mutations at specific TrkA autophosphorylation sites, we determined that mobility is controlled by multiple phosphorylation sites, but the SHC binding site at Y490 may be particularly important for ligand-induced immobilization of TrkA. Therefore, two mechanisms lead to NGF-induced immobilization of TrkAthe first resulting from autophosphorylation of TrkA and the second occurring through TrkA's association with gp75.