Inferring the Transcriptional Landscape of Bovine Skeletal Muscle by Integrating Co-Expression Networks

Background: Despite modern technologies and novel computational approaches, decoding causal transcriptional regulation remains challenging. This is particularly true for less well studied organisms and when only gene expression data is available. In muscle a small number of well characterised transcription factors are proposed to regulate development. Therefore, muscle appears to be a tractable system for proposing new computational approaches. Methodology/Principal Findings: Here we report a simple algorithm that asks "which transcriptional regulator has the highest average absolute co-expression correlation to the genes in a co-expression module?" It correctly infers a number of known causal regulators of fundamental biological processes, including cell cycle activity (E2F1), glycolysis (HLF), mitochondrial transcription (TFB2M), adipogenesis (PIAS1), neuronal development (TLX3), immune function (IRF1) and vasculogenesis (SOX17), within a skeletal muscle context. However, none of the canonical pro-myogenic transcription factors (MYOD1, MYOG, MYF5, MYF6 and MEF2C) were linked to muscle structural gene expression modules. Co-expression values were computed using developing bovine muscle from 60 days post conception (early foetal) to 30 months post natal (adulthood) for two breeds of cattle, in addition to a nutritional comparison with a third breed. A number of transcriptional landscapes were constructed and integrated into an always correlated landscape. One notable feature was a 'metabolic axis' formed from glycolysis genes at one end, nuclear-encoded mitochondrial protein genes at the other, and centrally tethered by mitochondrially-encoded mitochondrial protein genes. Conclusions/Significance: The new module-to-regulator algorithm complements our recently described Regulatory Impact Factor analysis. Together with a simple examination of a co-expression module's contents, these three gene expression approaches are starting to illuminate the in vivo transcriptional regulation of skeletal muscle development

Control of muscle mitochondria by insulin entails activation of Akt2-mtNOS pathway: implications for the metabolic syndrome.

BACKGROUND: In the metabolic syndrome with hyperinsulinemia, mitochondrial inhibition facilitates muscle fat and glycogen accumulation and accelerates its progression. In the last decade, nitric oxide (NO) emerged as a typical mitochondrial modulator by reversibly inhibiting citochrome oxidase and oxygen utilization. We wondered whether insulin-operated signaling pathways modulate mitochondrial respiration via NO, to alternatively release complete glucose oxidation to CO(2) and H(2)O or to drive glucose storage to glycogen. METHODOLOGY/PRINCIPAL FINDINGS: We illustrate here that NO produced by translocated nNOS (mtNOS) is the insulin-signaling molecule that controls mitochondrial oxygen utilization. We evoke a hyperinsulinemic-normoglycemic non-invasive clamp by subcutaneously injecting adult male rats with long-lasting human insulin glargine that remains stable in plasma by several hours. At a precise concentration, insulin increased phospho-Akt2 that translocates to mitochondria and determines in situ phosphorylation and substantial cooperative mtNOS activation (+4-8 fold, P<.05), high NO, and a lowering of mitochondrial oxygen uptake and resting metabolic rate (-25 to -60%, P<.05). Comparing in vivo insulin metabolic effects on gastrocnemius muscles by direct electroporation of siRNA nNOS or empty vector in the two legs of the same animal, confirmed that in the silenced muscles disrupted mtNOS allows higher oxygen uptake and complete (U-(14)C)-glucose utilization respect to normal mtNOS in the vector-treated ones (respectively 37+/-3 vs 10+/-1 micromolO(2)/h.g tissue and 13+/-1 vs 7.2+/-1 micromol (3)H(2)O/h.g tissue, P<.05), which reciprocally restricted glycogen-synthesis by a half. CONCLUSIONS/SIGNIFICANCE: These evidences show that after energy replenishment, insulin depresses mitochondrial respiration in skeletal muscle via NO which permits substrates to be deposited as macromolecules; at discrete hyperinsulinemia, persistent mtNOS activation could contribute to mitochondrial dysfunction with insulin resistance and obesity and therefore, to the progression of the metabolic syndrome

Abnormal mitochondrial fusion-fission balance contributes to the progression of experimental sepsis

Sepsis-associated multiple organ failure is a major cause of mortality characterized by a massive increase of reactive oxygen and nitrogen species (ROS/RNS) and mitochondrial dysfunction. Despite intensive research, determining events in the progression or reversal of the disease are incompletely understood. Herein, we studied two prototype sepsis models: endotoxemia and cecal ligation and puncture (CLP) - which showed very different lethality rates (2.5% and 67%, respectively) - , evaluated iNOS, ROS and respiratory chain activity, and investigated mitochondrial biogenesis and dynamics, as possible processes involved in sepsis outcome. Endotoxemia and CLP showed different iNOS, ROS/RNS, and complex activities time-courses. Moreover, these alterations reverted after 24-h endotoxemia but not after CLP. Mitochondrial biogenesis was not elicited during the first 24 h in either model but instead, 50% mtDNA depletion was observed. Mitochondrial fusion and fission were evaluated using real-time PCR of mitofusin-2 (Mfn2), dynamin-related protein-1 (Drp1), and using electron microscopy. During endotoxemia, we observed a decrease of Mfn2-mRNA levels at 4-6 h, and an increase of mitochondrial fragmentation at 6 h. These parameters reverted at 24 h. In contrast, CLP showed not only decreased Mfn2-mRNA levels at 12-18 h but also increased Drp1-mRNA levels at 4 h, and enhanced and sustained mitochondrial fragmentation. The in vivo pretreatment with mdivi-1 (Drp1 inhibitor) significantly attenuated mitochondrial dysfunction and apoptosis in CLP. Therefore, abnormal fusion-to-fission balance, probably evoked by ROS/RNS secondary to iNOS induction, contributes to the progression of sepsis. Pharmacological targeting of Drp1 may be a potential novel therapeutic tool for sepsis.Fil: Gonzalez, Analia Silvia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Elguero, María Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Finocchietto, P.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Holod, Silvia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Romorini, Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Miriuka, Santiago Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Peralta, J. G.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Poderoso, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Carreras, Maria Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentin

Cardiac-specific overexpression of thioredoxin 1 attenuates mitochondrial and myocardial dysfunction in septic mice

Sepsis-induced myocardial dysfunction is associated with increased oxidative stress and mitochondrial dysfunction. Current evidence suggests a protective role of thioredoxin-1 (Trx1) in the pathogenesis of cardiovascular diseases. However, it is unknown yet a putative role of Trx1 in sepsis-induced myocardial dysfunction, in which oxidative stress is an underlying cause. Transgenic male mice with Trx1 cardiac-specific overexpression (Trx1-Tg) and its wild-type control (wt) were subjected to cecal ligation and puncture or sham surgery. After 6, 18, and 24 h, cardiac contractility, antioxidant enzymes, protein oxidation, and mitochondrial function were evaluated. Trx1 overexpression improved the average life expectancy (Trx1-Tg: 36, wt: 28 h; p = 0.0204). Sepsis induced a decrease in left ventricular developed pressure in both groups, while the contractile reserve, estimated as the response to β-adrenergic stimulus, was higher in Trx1-Tg in relation to wt, after 6 h of the procedure. Trx1 overexpression attenuated complex I inhibition, protein carbonylation, and loss of membrane potential, and preserved Mn superoxide dismutase activity at 24 h. Ultrastructural alterations in mitochondrial cristae were accompanied by reduced optic atrophy 1 (OPA1) fusion protein, and activation of dynamin-related protein 1 (Drp1) (fission protein) in wt mice at 24 h, suggesting mitochondrial fusion/fission imbalance. PGC-1α gene expression showed a 2.5-fold increase in Trx1-Tg at 24 h, suggesting mitochondrial biogenesis induction. Autophagy, demonstrated by electron microscopy and increased LC3-II/LC3-I ratio, was observed earlier in Trx1-Tg. In conclusion, Trx1 overexpression extends antioxidant protection, attenuates mitochondrial damage, and activates mitochondrial turnover (mitophagy and biogenesis), preserves contractile reserve and prolongs survival during sepsis.Fil: Sánchez Villamil, Juana P.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: D'Anunzio, Verónica. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Finocchietto, Paola Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Holod, Silvia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Bioquímica Clínica; ArgentinaFil: Rebagliati, Ines Rosa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Pérez, Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Peralta, Jorge G.. Universidad de Buenos Aires. Facultad de Medicina; ArgentinaFil: Gelpi, Ricardo Jorge. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; ArgentinaFil: Poderoso, Juan José. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; ArgentinaFil: Carreras, Maria Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Bioquímica Clínica; Argentin