Proto-oncogenic H-Ras, K-Ras, and N-Ras are involved in muscle differentiation via phosphatidylinositol 3-kinase

Abstract

Oncogenic H-Ras G12V and its variants have been shown to inhibit muscle differentiation. However, the role of proto-oncogenic Ras (c-Ras) in muscle differentiation remains unclear. The active GTP-bound form of Ras has been known to associate with diverse effectors including Raf, phosphatidylinositol 3-kinase (PI3K), Ral-GDS, and other molecules to transmit downstream signals. We hypothesize that c-Ras may stimulate muscle differentiation by selectively activating PI3K, an important mediator for muscle differentiation. In our experiments, inhibition of c-Ras by farnesyltransferase inhibitors and a dominant negative form of H-Ras (Ras S17N) suppressed muscle differentiation. Consistently, individual knockdown of H-Ras, K-Ras, and N-Ras by siRNAs all blocked muscle differentiation. Interestingly, we found that c-Ras preferentially interacts with PI3K rather than its major binding partner c-Raf, during myogenic differentiation, with total c-Ras activity remaining unchanged. PI3K and its downstream myogenic pathway, the Nox2/NF-κB/inducible nitric oxide synthase (iNOS) pathway, were found to be suppressed by inhibition of c-Ras activity during differentiation. Furthermore, expression of a constitutively active form of PI3K completely rescued the differentiation block and reactivated the Nox2/NF-κB/iNOS pathway in c-Ras-inhibited cells. On the basis of our results, we conclude that contrary to oncogenic Ras, proto-oncogenic H-Ras, K-Ras, and N-Ras are directly involved in the promotion of muscle differentiation via PI3K and its downstream signaling pathways. In addition, PI3K pathway activation is associated with a concurrent suppression of the otherwise predominantly activated Raf/ Mek/Erk pathway.

Manumycin A corrects aberrant splicing of Clcn1 in myotonic dystrophy type 1 (DM1) mice

“…A recent study revealed that oncogenic mutated H-Ras G12V inhibits muscle differentiation44. If H-Ras is involved in DM pathology, it may help us to better understand DM1.…”

Immunostimulating activity of maysin isolated from corn silk in murine RAW 264.7 macrophages

“…Nuclear extracts were obtained, according to a previous report, with minor modifications (25). Briefly, confluent cells in 10-cm-diame ter dishes were resuspended in 200 μl of buffer (10 mM HEPES, pH 7.9, 10 mM KCl, 1 mM DTT, 0.5 mM EDTA, 0.5 mM PMSF), and lysed with 12.5 μl of 10% NP-40.…”

Mitochondria-targeted Antioxidants Protect Pancreatic β-cells against Oxidative Stress and Improve Insulin Secretion in Glucotoxicity and Glucolipotoxicity

“…The intracellular ROS and mitochondrial membrane potential were measured using flow cytometry as previously described [29,30,32]. In brief, equal numbers of RINm5F cells were seeded in a 6-well microplate and grown to 80% confluency in RPMI medium.…”