Given that receptor PTKs are thought to be activated by extracellular cues and transduce such stimuli into cytoplasmic signaling, this observation is surprising. However, downregulation of Dok-7 using RNA interference technique demonstrated that Dok-7 is required for activation of MuSK at least in cultured myotubes (14). Furthermore, the RNA interference experiments revealed that even neural Agrin requires Dok-7 to activate MuSK in myotubes (14). Indeed, E18.5 embryos of mice lacking Dok-7 do not form AChR clusters nor NMJs in the diaphragm muscles as Inhibitors,research,lifescience,medical was observed in MuSK-deficient mice (14, 17). It is of note that both mutant mice showed abnormal extension of motor nerve

axons at the medial area of the ABT-888 datasheet skeletal muscle possibly due to the lack of retrograde signaling from the postsynaptic apparatus (14, 17). Together, these data indicate that Dok-7 is a cytoplasmic activator of MuSK essential for MuSK-dependent postsynaptic Inhibitors,research,lifescience,medical specialization of NMJ (Fig. ​(Fig.11). Figure 1 A

greatly simplified model of the Dok-7/MuSK pathway Inhibitors,research,lifescience,medical for postsynaptic specialization of the mammalian NMJ. Dok-7 can activate MuSK and induce Rapsyn-dependent AChR clustering even in the absence of neural Agrin; however MuSK requires both Dok-7 and Agrin … As previously stated, a skeletal muscle-intrinsic activator of MuSK was predicted due to existence of i) aneural, but MuSK-dependent, clustering of AChRs in mouse embryos; and ii) neuromuscular synapse Inhibitors,research,lifescience,medical formation in mice lacking both Agrin and CHAT (5–7). Although there is no definitive proof that Dok-7 is this muscle-intrinsic activator of MuSK, there is supporting evidence: i) mice lacking Dok-7 or MuSK form no AChR clusters while those lacking Agrin can form aneural, MuSK-dependent AChR clusters (14, 17); ii) Dok-7 transcripts were preferentially expressed in the central region of the diaphragm

muscle of mouse E14.5 embryos, where the aneural, MuSK-dependent Inhibitors,research,lifescience,medical AChR clusters normally form (14). However, as we will discuss later, if Dok-7 plays a role as an essential signaling molecule downstream of MuSK, the same defects could be observed in Dok-7-deficient mice. Therefore, careful examination of kinase activity of MuSK in the skeletal muscle during embryogenesis of Dok-7-deficient mice would be important. The Dok-7/MuSK pathway How does a cytoplasmic adaptor-like protein Dok-7 activate a receptor PTK MuSK? Although the definitive Levetiracetam answer awaits further studies, several interesting observations have been found (14). In heterologous cells, which do not express Dok-7 nor MuSK, the forced expression of these two proteins resulted in robust activation of MuSK. In addition, when ectopically expressed in heterologous cells, these proteins form a stable complex that requires the intact PTB domain of Dok-7 and its target motif encompassing Tyr-553 in the juxtamembrane region of MuSK.