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 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 DAPT secretase datasheet 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 most 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.