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Nitric Oxide Regulates Skeletal Muscle Fatigue, Fiber Type, Microtubule Organization, and Mitochondrial ATP Synthesis Efficiency Through cGMP-Dependent Mechanisms.

ABSTRACT: Skeletal muscle nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathways are impaired in Duchenne and Becker muscular dystrophy partly because of reduced nNOS? and soluble guanylate cyclase (GC) activity. However, GC function and the consequences of reduced GC activity in skeletal muscle are unknown. In this study, we explore the functions of GC and NO-cGMP signaling in skeletal muscle.GC1, but not GC2, expression was higher in oxidative than glycolytic muscles. GC1 was found in a complex with nNOS? and targeted to nNOS compartments at the Golgi complex and neuromuscular junction. Baseline GC activity and GC agonist responsiveness was reduced in the absence of nNOS. Structural analyses revealed aberrant microtubule directionality in GC1-/- muscle. Functional analyses of GC1-/- muscles revealed reduced fatigue resistance and postexercise force recovery that were not due to shifts in type IIA-IIX fiber balance. Force deficits in GC1-/- muscles were also not driven by defects in resting mitochondrial adenosine triphosphate (ATP) synthesis. However, increasing muscle cGMP with sildenafil decreased ATP synthesis efficiency and capacity, without impacting mitochondrial content or ultrastructure.GC may represent a new target for alleviating muscle fatigue and that NO-cGMP signaling may play important roles in muscle structure, contractility, and bioenergetics.These findings suggest that GC activity is nNOS dependent and that muscle-specific control of GC expression and differential GC targeting may facilitate NO-cGMP signaling diversity. They suggest that nNOS regulates muscle fiber type, microtubule organization, fatigability, and postexercise force recovery partly through GC1 and suggest that NO-cGMP pathways may modulate mitochondrial ATP synthesis efficiency. Antioxid. Redox Signal. 26, 966-985.

SUBMITTER: Moon Y

PROVIDER: S-EPMC5467110 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Nitric Oxide Regulates Skeletal Muscle Fatigue, Fiber Type, Microtubule Organization, and Mitochondrial ATP Synthesis Efficiency Through cGMP-Dependent Mechanisms.

Moon Younghye Y Balke Jordan E JE Madorma Derik D Siegel Michael P MP Knowels Gary G Brouckaert Peter P Buys Emmanuel S ES Marcinek David J DJ Percival Justin M JM

Antioxidants & redox signaling 20160817 17

<h4>Aim</h4>Skeletal muscle nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathways are impaired in Duchenne and Becker muscular dystrophy partly because of reduced nNOSμ and soluble guanylate cyclase (GC) activity. However, GC function and the consequences of reduced GC activity in skeletal muscle are unknown. In this study, we explore the functions of GC and NO-cGMP signaling in skeletal muscle.<h4>Results</h4>GC1, but not GC2, expression was higher in oxidative than glycolytic muscles. ...[more]

PMID: 27393340

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Project description:BackgroundRegulatory functions of nitric oxide (NO*) that bypass the second messenger cGMP are incompletely understood. Here, cGMP-independent effects of NO* on gene expression were globally examined in U937 cells, a human monoblastoid line that constitutively lacks soluble guanylate cyclase. Differentiated U937 cells (>80% in G0/G1) were exposed to S-nitrosoglutathione, a NO* donor, or glutathione alone (control) for 6 h without or with dibutyryl-cAMP (Bt2cAMP), and then harvested to extract total RNA for microarray analysis. Bt2cAMP was used to block signaling attributable to NO*-induced decreases in cAMP.ResultsNO* regulated 110 transcripts that annotated disproportionately to the cell cycle and cell proliferation (47/110, 43%) and more frequently than expected contained AU-rich, post-transcriptional regulatory elements (ARE). Bt2cAMP regulated 106 genes; cell cycle gene enrichment did not reach significance. Like NO*, Bt2cAMP was associated with ARE-containing transcripts. A comparison of NO* and Bt2cAMP effects showed that NO* regulation of cell cycle genes was independent of its ability to interfere with cAMP signaling. Cell cycle genes induced by NO* annotated to G1/S (7/8) and included E2F1 and p21/Waf1/Cip1; 6 of these 7 were E2F target genes involved in G1/S transition. Repressed genes were G2/M associated (24/27); 8 of 27 were known targets of p21. E2F1 mRNA and protein were increased by NO*, as was E2F1 binding to E2F promoter elements. NO* activated p38 MAPK, stabilizing p21 mRNA (an ARE-containing transcript) and increasing p21 protein; this increased protein binding to CDE/CHR promoter sites of p21 target genes, repressing key G2/M phase genes, and increasing the proportion of cells in G2/M.ConclusionNO* coordinates a highly integrated program of cell cycle arrest that regulates a large number of genes, but does not require signaling through cGMP. In humans, antiproliferative effects of NO* may rely substantially on cGMP-independent mechanisms. Stress kinase signaling and alterations in mRNA stability appear to be major pathways by which NO* regulates the transcriptome.

Project description:Nitric oxide activates guanylate cyclase to form cGMP, comprising a signalling system that is believed to be a distinct mechanism for increasing glucose transport and metabolism in skeletal muscle. The effects of a selective cGMP phosphodiesterase inhibitor, zaprinast, on basal glucose utilization was investigated in incubated rat soleus muscle preparations isolated from both insulin-sensitive (lean Zucker; Fa/?) and insulin-resistant (obese Zucker; fa/fa) rats. Zaprinast at 27 microM significantly increased cGMP levels in incubated soleus muscle isolated from lean, but not obese, Zucker rats. Muscles were incubated with 14C-labelled glucose and various concentrations of zaprinast (3, 27 and 243 microM). Zaprinast (at 27 and 243 microM) significantly increased rates of net and 14C-labelled lactate release and of glycogen synthesis in lean Zucker rat soleus muscle; glucose oxidation was also increased by 27 microM zaprinast. In addition, regardless of concentration, the phosphodiesterase inhibitor failed to increase any aspect of 14C-labelled glucose utilization in soleus muscles isolated from obese Zucker rats. The maximal activity of nitric oxide synthase (NOS) was significantly decreased in insulin-resistant obese Zucker muscles. Thus the lack of effect of zaprinast in insulin-resistant skeletal muscle is consistent with decreased NOS activity. To test whether there is a defect in insulin-resistant skeletal muscle for endogenous activation of guanylate cyclase, soleus muscles were isolated from both insulin-sensitive and insulin-resistant Zucker rats and incubated with various concentrations of the NO donor sodium nitroprusside (SNP; 0.1, 1, 5 and 15 mM). SNP significantly increased rates of net and 14C-labelled lactate release, as well as glucose oxidation in muscles isolated from both insulin-sensitive and insulin-resistant rats. A decreased response to SNP was observed in the dose-dependent generation of cGMP within isolated soleus muscles from insulin-resistant rats. A possible link between impaired NO/cGMP signalling and abnormal glucose utilization by skeletal muscle is discussed.

Project description:Cyclic nucleotide-gated (CNG) channels are a family of ion channels activated by the binding of cyclic nucleotides. Endogenous channels have been used to measure cyclic nucleotide signals in photoreceptor outer segments and olfactory cilia for decades. Here we have investigated the subcellular localization of cGMP signals by monitoring CNG channel activity in response to agonists that activate either particulate or soluble guanylyl cyclase. CNG channels were heterologously expressed in either human embryonic kidney (HEK)-293 cells that stably overexpress a particulate guanylyl cyclase (HEK-NPRA cells), or cultured vascular smooth muscle cells (VSMCs). Atrial natriuretic peptide (ANP) was used to activate the particulate guanylyl cyclase and the nitric oxide donor S-nitroso-n-acetylpenicillamine (SNAP) was used to activate the soluble guanylyl cyclase. CNG channel activity was monitored by measuring Ca2+ or Mn2+ influx through the channels using the fluorescent dye, fura-2. We found that in HEK-NPRA cells, ANP-induced increases in cGMP levels activated CNG channels in a dose-dependent manner (0.05-10 nM), whereas SNAP (0.01-100 microM) induced increases in cGMP levels triggered little or no activation of CNG channels (P < 0.01). After pretreatment with 100 microM 3-isobutyl-1-methylxanthine (IBMX), a nonspecific phosphodiesterase inhibitor, ANP-induced Mn2+ influx through CNG channels was significantly enhanced, while SNAP-induced Mn2+ influx remained small. In contrast, we found that in the presence of IBMX, both 1 nM ANP and 100 microM SNAP triggered similar increases in total cGMP levels. We next sought to determine if cGMP signals are compartmentalized in VSMCs, which endogenously express particulate and soluble guanylyl cyclase. We found that 10 nM ANP induced activation of CNG channels more readily than 100 muM SNAP; whereas 100 microM SNAP triggered higher levels of total cellular cGMP accumulation. These results suggest that cGMP signals are spatially segregated within cells, and that the functional compartmentalization of cGMP signals may underlie the unique actions of ANP and nitric oxide.

Dataset Information

Nitric Oxide Regulates Skeletal Muscle Fatigue, Fiber Type, Microtubule Organization, and Mitochondrial ATP Synthesis Efficiency Through cGMP-Dependent Mechanisms.

Publications

Nitric Oxide Regulates Skeletal Muscle Fatigue, Fiber Type, Microtubule Organization, and Mitochondrial ATP Synthesis Efficiency Through cGMP-Dependent Mechanisms.

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