Downstream of tyrosine kinase/docking protein 6, as a novel substrate of tropomyosin-related kinase C receptor, is involved in neurotrophin 3-mediated neurite outgrowth in mouse cortex neurons.
BACKGROUND
The downstream of tyrosine kinase/docking protein (Dok) adaptor protein family has seven members, Dok1 to Dok7, that act as substrates of multiple receptor tyrosine kinase and non-receptor tyrosine kinase. The tropomyosin-related kinase (Trk) receptor family, which has three members (TrkA, TrkB and TrkC), are receptor tyrosine kinases that play pivotal roles in many stages of nervous system development, such as differentiation, migration, axon and dendrite projection and neuron patterning. Upon related neurotrophin growth factor stimulation, dimerisation and autophosphorylation of Trk receptors can occur, recruiting adaptor proteins to mediate signal transduction.


RESULTS
In this report, by using yeast two-hybrid assays, glutathione S-transferase (GST) precipitation assays and coimmunoprecipitation (Co-IP) experiments, we demonstrate that Dok6 selectively binds to the NPQY motif of TrkC through its phosphotyrosine-binding (PTB) domain in a kinase activity-dependent manner. We further confirmed their interaction by coimmunoprecipitation and colocalisation in E18.5 mouse cortex neurons, which provided more in vivo evidence. Next, we demonstrated that Dok6 is involved in neurite outgrowth in mouse cortex neurons via the RNAi method. Knockdown of Dok6 decreased neurite outgrowth in cortical neurons upon neurotrophin 3 (NT-3) stimulation.


CONCLUSIONS
We conclude that Dok6 interacts with the NPQY motif of the TrkC receptor through its PTB domain in a kinase activity-dependent manner, and works as a novel substrate of the TrkC receptor involved in NT-3-mediated neurite outgrowth in mouse cortex neurons.
Li
Wei qi
Wq
National Key Laboratory of Medical Molecular Biology, School of Basic Medicine, Peking Union Medical College, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, Beijing, China.
Shi
Lei
L
You
Yuan gang
Yg
Gong
Yan hua
Yh
Yin
Bin
B
Yuan
Jian gang
Jg
Peng
Xiao zhong
Xz
D048868:Adaptor Proteins, Signal Transducing / Q000032:analysis / Q000378:metabolism*; D020816:Amino Acid Motifs; D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000648:ultrastructure; D005982:Glutathione Transferase / Q000378:metabolism*; D006801:Humans; D051379:Mice; D016501:Neurites / Q000378:metabolism*; D020933:Neurotrophin 3 / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D020812:Receptor, trkC / Q000032:analysis / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D048868:Adaptor Proteins, Signal Transducing; C553041:DOK6 protein, mouse; D020933:Neurotrophin 3; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases; D020812:Receptor, trkC