Myotubularin Regulates Akt-dependent Survival Signaling via Phosphatidylinositol 3-Phosphate

Abstract

Myotubularin is a 3-phosphoinositide phosphatase that is mutated in X-linked myotubular myopathy, a severe neonatal disorder in which skeletal muscle development and/or regeneration is impaired. In this report we provide evidence that siRNA-mediated silencing of myotubularin expression markedly inhibits growth factor-stimulated Akt phosphorylation, leading to activation of caspase-dependent pro-apoptotic signaling in HeLa cells and primary human skeletal muscle myotubes. Myotubularin silencing also inhibits Akt-dependent signaling through the mammalian target of rapamycin complex 1 as assessed by p70 S6-kinase and 4E-BP1 phosphorylation. Similarly, phosphorylation of FoxO transcription factors is also significantly reduced in myotubularin-deficient cells. Our data further suggest that inhibition of Akt activation and downstream survival signaling in myotubularin-deficient cells is caused by accumulation of the MTMR substrate lipid phosphatidylinositol 3-phosphate generated from the type II phosphatidylinositol 3-kinase PIK3C2B. Our findings are significant because they suggest that myotubularin regulates Akt activation via a cellular pool of phosphatidylinositol 3-phosphate that is distinct from that generated by the type III phosphatidylinositol 3-kinase hVps34. Because impaired Akt signaling has been tightly linked to skeletal muscle atrophy, we hypothesize that loss of Akt-dependent growth/survival cues due to impaired myotubularin function may be a critical factor underlying the severe skeletal muscle atrophy characteristic of muscle fibers in patients with X-linked myotubular myopathy.Myotubularin is the archetypical member of a family of phosphoinositide 3-phosphatases. Mutations in the myotubularin gene are causative for X-linked recessive myotubular myopathy (XLMTM), 4 also known as "centronuclear myopathy." XLMTM is a severe neonatal disorder in which the maturation and/or regeneration of skeletal muscle fibers is compromised (1, 2). Myofibers from affected individuals display abnormal centrally located nuclei and are severely atrophic. Mice in which the myotubularin gene has been deleted exhibit a skeletal muscle phenotype consistent with that of XLMTM disease. Like XLMTM patients, early stages of myogenesis in these mice appear to occur normally, and it has been suggested that skeletal muscle maintenance or regeneration is affected rather than early myogenic differentiation (3). Mutations in two other MTMR family proteins, MTMR2 and MTMR13, are causative for the demyelinating peripheral neuropathy type 4B Charcot-Marie-Tooth disease (4 -6).The myotubularin-related protein (MTMR) family consists of 14 proteins including 8 catalytically active lipid phosphatases and 6 forms that are inactive due to germ-line substitutions of essential catalytic residues (7). Active MTMRs specifically remove the 3-phosphate moiety from phosphatidylinositol 3-phosphate and phosphatidylinositol 3,5-bisphosphate (PI(3,5)P 2 ), generating phosphatidylinositol (PI) and phosphatidylinositol 5-phosphate (PI(5)P), respectively (...

Myotubularin-related protein 7 inhibits insulin signaling in colorectal cancer

“…Inhibition of the AKT pathway by MTMs has been reported for T lymphocytes [23] and myotubes [24, 25] in mice and zebrafish [22]. Thus, our data may contribute to redefine MTMs from “survival” [19, 20, 23, 26] towards “tumor suppressor” phosphatases similar to PTEN [27]. MTMR7 blocked RTK signaling in KRAS -WT and KRAS -mutated cells, and there was no correlation between MTMR7 loss and KRAS -mutations in CRC patients.…”

“…MTMR7 decreased phosphorylation of ERK1/2 by MEK1/2 on 202/185TEY204/187 and of AKT on S473/T308 residues. Hence, MTMR7 did not block single mTORC1/2 complexes, as described for MTM1 [19, 20] or MTMR3 [21], what would have abrogated phosphorylation exclusively on either SER473, which is part of the AKT-mTORC2 feed forward loop, or THR308, which is targeted by PI3K-PDK1. Instead, MTMR7 inhibited upstream components in these pathways resulting in a decreased production of the lipid second messenger PI(3)P. MTMR8 binds PI3K thereby reducing PI3K activity [22].…”

An Introduction to Phosphoinositides

“…For instance, down-regulation of MTMs in C. elegans is able to rescue the phenotype of a Vps34 null mutant (Xue et al 2003). Similarly, downregulation of PI3K-C2b specifically rescues the inhibition of Akt phosphorylation induced by down-regulation of MTM1 (Razidlo et al 2011). The mechanism of activation of class II PI3Ks is still not clear although data suggest that it may involve translocation of the enzymes to the plasma membrane (Maffucci et al 2005;Falasca and Maffucci 2012).…”

PIK3C2B inhibition improves function and prolongs survival in myotubular myopathy animal models

“…On the basis of our discovery of increased PI3P levels in MTM, and observations from cell culture (15) and invertebrate studies (16,17) showing reciprocal interplay between MTM1 and PI3P-synthesizing kinases, we hypothesized that lowering PI3P levels may prevent MTM disease progression. We therefore examined the kinases responsible for generating PI3P (i.e., the class II and class III PI3Ks).…”