Neutral sphingomyelinase-3 mediates TNF-stimulated oxidant activity in skeletal muscle

Abstract

AimsSphingolipid and oxidant signaling affect glucose uptake, atrophy, and force production of skeletal muscle similarly and both are stimulated by tumor necrosis factor (TNF), suggesting a connection between systems. Sphingolipid signaling is initiated by neutral sphingomyelinase (nSMase), a family of agonist-activated effector enzymes. Northern blot analyses suggest that nSMase3 may be a striated muscle-specific nSMase. The present study tested the hypothesis that nSMase3 protein is expressed in skeletal muscle and functions to regulate TNF-stimulated oxidant production.ResultsWe demonstrate constitutive nSMase activity in skeletal muscles of healthy mice and humans and in differentiated C2C12 myotubes. nSMase3 (Smpd4 gene) mRNA is highly expressed in muscle. An nSMase3 protein doublet (88 and 85 kD) is derived from alternative mRNA splicing of exon 11. The proteins partition differently. The full-length 88 kD isoform (nSMase3a) fractionates with membrane proteins that are resistant to detergent extraction; the 85 kD isoform lacking exon 11 (nSMase3b) is more readily extracted and fractionates with detergent soluble membrane proteins; neither variant is detected in the cytosol. By immunofluorescence microscopy, nSMase3 resides in both internal and sarcolemmal membranes. Finally, myotube nSMase activity and cytosolic oxidant activity are stimulated by TNF. Both if these responses are inhibited by nSMase3 knockdown.InnovationThese findings identify nSMase3 as an intermediate that links TNF receptor activation, sphingolipid signaling, and skeletal muscle oxidant production.ConclusionOur data show that nSMase3 acts as a signaling nSMase in skeletal muscle that is essential for TNF-stimulated oxidant activity.

Inhibitors of the sphingomyelin cycle: Sphingomyelin synthases and sphingomyelinases

“…A recent report showed that this enzyme could play a role in DNA damage responses in human tumorigenesis (Corcoran et al, 2008) and in response of oxidative stress in skeletal muscle (Moylan et al, 2014). Clearly additional studies are needed to address the role of nSMase3 in cancer and in sphingolipid biology.…”

Skeletal Muscle Alterations Are Exacerbated in Heart Failure With Reduced Compared With Preserved Ejection Fraction

“…32 Similar to our findings, TNF-α is increased in patients with HFrEF [33][34][35] and is a key factor regulating muscle mass, 36 which can activate the expression of atrogenes such as MuRF1 and MafBx 37 while suppressing anabolic factors such as IGF-1, 38 while also further mediating oxidative stress. 39 In addition, TNF-α can downregulate PGC-1α expression and thus impair mitochondrial function. 40 That we too found such alterations in our HFrEF animals simultaneous with increased plasma TNF-α levels strongly suggests a causal role of this cytokine in mediating skeletal muscle alterations between HF cohorts-a suggestion further supported by our HFpEF animals having unchanged plasma concentrations of TNF-α alongside fewer skeletal muscle alterations.…”

Loss of SMPD4 Causes a Developmental Disorder Characterized by Microcephaly and Congenital Arthrogryposis

“…Although nSMase-3 (from now on indicated as SMPD4), encoded by SMPD4, has no significant sequence homology with other sphingomyelinases, some papers reported its catalytic activity and described regulation of its expression by tumor necrosis factor and p53, implicating this protein in response to cellular stress. [4][5][6] SMPD4 is a 866 amino acid long protein with a protein-protein interaction domain and a C-terminal transmembrane domain close to the putative enzyme active site, which includes a potential N-glycosylation site. 5,6 Subcellular localization experiments have shown the presence of SMPD4 at the endoplasmic reticulum (ER) and the Golgi apparatus.…”

“…[4][5][6] SMPD4 is a 866 amino acid long protein with a protein-protein interaction domain and a C-terminal transmembrane domain close to the putative enzyme active site, which includes a potential N-glycosylation site. 5,6 Subcellular localization experiments have shown the presence of SMPD4 at the endoplasmic reticulum (ER) and the Golgi apparatus. In independent studies, proteomics analysis of nuclear fractions detected SMPD4 at the nuclear envelope.…”