Abstract
Myotubularin and related proteins constitute a large and highly conserved family possessing phosphoinositide 3-phosphatase activity, although not all members possess this activity. This family contains a conserved region called the GRAM domain that is found in a variety of proteins associated with membrane-coupled processes and signal transduction. Mutations of myotubularin are found in X-linked myotubular myopathy, a severe muscle disease. Mutations in the GRAM domain are responsible for this condition, suggesting crucial roles for this region. Here, we show that the GRAM domain of myotubularin binds to phosphoinositide with the highest affinity to phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P(2)). In patients with myotubular myopathy, mutations in the myotubularin GRAM domain eliminate this binding, indicating that the PtdIns(3,5)P(2) binding ability of the GRAM (glucosyltransferases, Rablike GTPase activators and myotubularin) domain is crucial for the functions of myotubularin in vivo. Stimulation of epidermal growth factor recruits myotubularin to the late endosomal compartment in a manner dependent on the phosphoinositide binding. Overexpression of myotubularin inhibits epidermal growth factor receptor trafficking from late endosome to lysosome and induces the large endosomal vacuoles. Thus, our data suggest that myotubularin phosphatase physiologically functions in late endosomal trafficking and vacuolar morphology through interaction with PtdIns(3,5)P(2).
Highlights
Myotubularin and related proteins constitute a large and highly conserved family possessing phosphoinositide 3-phosphatase activity, not all members possess this activity
We further demonstrate that overexpression of myotubularin inhibits epidermal growth factor receptor (EGFR) trafficking from the late endosome to the lysosome and induces large endosomal vacuoles
The GRAM Domain Binds to PtdIns(3,5)P2—We examined the possibility that the GRAM domain of myotubularin and related proteins could bind to phosphoinositides
Summary
Myotubularin and related proteins constitute a large and highly conserved family possessing phosphoinositide 3-phosphatase activity, not all members possess this activity. This binding is abolished by point mutations found in the GRAM domain of myotubularin in myopathy patients, indicating that interaction with PtdIns (3,5)P2 is essential for full function of this phosphatase.
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