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In cell biology, ribosomal RNA (rRNA) 2′O-methyl (2′-O-Me) is the most prevalent posttranscriptional chemical modification contributing to ribosome heterogeneity. The modification involves a family of small nucleolar RNAs (snoRNAs) and is specified by box C/D snoRNAs (SNORDs). Given the importance of ribosome biogenesis for skeletal muscle growth, we asked if rRNA 2′-O-Me in nascent ribosomes synthesized in response to a growth stimulus is an unrecognized mode of ribosome heterogeneity in muscle. To determine the pattern and dynamics of 2′-O-Me rRNA, we used a sequencing-based profiling method called RiboMeth-seq (RMS). We applied this method to tissue-derived rRNA of skeletal muscle and rRNA specifically from the muscle fiber using an inducible myofiber-specific RiboTag mouse in sedentary and mechanically overloaded conditions. These analyses were complemented by myonuclear-specific small RNA sequencing to profile SNORDs and link the rRNA epitranscriptome to known regulatory elements generated within the muscle fiber. We demonstrate for the first time that mechanical overload of skeletal muscle 1) induces decreased 2′-O-Me at a subset of skeletal muscle rRNA and 2) alters the SNORD profile in isolated myonuclei. These findings point to a transient diversification of the ribosome pool via 2′-O-Me during growth and adaptation in skeletal muscle. These findings suggest changes in ribosome heterogeneity at the 2′-O-Me level during muscle hypertrophy and lay the foundation for studies investigating the functional implications of these newly identified "growth-induced" ribosomes. NEW & NOTEWORTHY: Ribosomal RNAs (rRNAs) are posttranscriptionally modified by 2′O-methyl (2′-O-Me). This study applied RiboMeth-seq (RMS) to detect changes in 2′-O-Me levels during skeletal muscle hypertrophy, uncovering transient diversification of the ribosome pool in skeletal muscle fibers. This work implies a role for ribosome heterogeneity in skeletal muscle growth and adaptation. [ABSTRACT FROM AUTHOR]

The central dogma of molecular biology dictates the general flow of molecular information from DNA that leads to a functional cellular outcome. In skeletal muscle fibers, the extent to which global myonuclear transcriptional alterations, accounting for epigenetic and post-transcriptional influences, contribute to an adaptive stress response is not clearly defined. In this investigation, we leveraged an integrated analysis of the myonucleus-specific DNA methylome and transcriptome, as well as myonuclear small RNA profiling to molecularly define the early phase of skeletal muscle fiber hypertrophy. The analysis of myonucleus-specific mature microRNA and other small RNA species provides new directions for exploring muscle adaptation and complemented the methylation and transcriptional information. Our integrated multi-omics interrogation revealed a coordinated myonuclear molecular landscape during muscle loading that coincides with an acute and rapid reduction of oxidative metabolism. This response may favor a biosynthesis-oriented metabolic program that supports rapid hypertrophic growth. Graphical Abstract [ABSTRACT FROM AUTHOR]

Background: Prolonged exposure to toxic heavy metals leads to deleterious health outcomes including kidney injury. Metal exposure occurs through both environmental pathways including contamination of drinking water sources and from occupational hazards, including the military-unique risks from battlefield injuries resulting in retained metal fragments from bullets and blast debris. One of the key challenges to mitigate health effects in these scenarios is to detect early insult to target organs, such as the kidney, before irreversible damage occurs. Methods: High-throughput transcriptomics (HTT) has been recently demonstrated to have high sensitivity and specificity as a rapid and cost-effective assay for detecting tissue toxicity. To better understand the molecular signature of early kidney damage, we performed RNA sequencing (RNA-seq) on renal tissue using a rat model of soft tissue-embedded metal exposure. We then performed small RNA-seq analysis on serum samples from the same animals to identify potential miRNA biomarkers of kidney damage. Results: We found that metals, especially lead and depleted uranium, induce oxidative damage that mainly cause dysregulated mitochondrial gene expression. Utilizing publicly available single-cell RNA-seq datasets, we demonstrate that deep learning-based cell type decomposition effectively identified cells within the kidney that were affected by metal exposure. By combining random forest feature selection and statistical methods, we further identify miRNA-423 as a promising early systemic marker of kidney injury. Conclusion: Our data suggest that combining HTT and deep learning is a promising approach for identifying cell injury in kidney tissue. We propose miRNA-423 as a potential serum biomarker for early detection of kidney injury. [ABSTRACT FROM AUTHOR]

Background: Skeletal muscle (SkM) is a large, secretory organ that produces and releases myokines that can have autocrine, paracrine, and endocrine effects. Whether extracellular vesicles (EVs) also play a role in the SkM adaptive response and ability to communicate with other tissues is not well understood. The purpose of this study was to investigate EV biogenesis factors, marker expression, and localization across cell types in the skeletal muscle. We also aimed to investigate whether EV concentrations are altered by disuse atrophy. Methods: To identify the potential markers of SkM-derived EVs, EVs were isolated from rat serum using density gradient ultracentrifugation, followed by fluorescence correlation spectroscopy measurements or qPCR. Single-cell RNA sequencing (scRNA-seq) data from rat SkM were analyzed to assess the EV biogenesis factor expression, and cellular localization of tetraspanins was investigated by immunohistochemistry. Finally, to assess the effects of mechanical unloading on EV expression in vivo, EV concentrations were measured in the serum by nanoparticle tracking analysis in both a rat and human model of disuse. Results: In this study, we show that the widely used markers of SkM-derived EVs, α-sarcoglycan and miR-1, are undetectable in serum EVs. We also found that EV biogenesis factors, including the tetraspanins CD63, CD9, and CD81, are expressed by a variety of cell types in SkM. SkM sections showed very low detection of CD63, CD9, and CD81 in myofibers and instead accumulation within the interstitial space. Furthermore, although there were no differences in serum EV concentrations following hindlimb suspension in rats, serum EV concentrations were elevated in human subjects after bed rest. Conclusions: Our findings provide insight into the distribution and localization of EVs in SkM and demonstrate the importance of methodological guidelines in SkM EV research. [ABSTRACT FROM AUTHOR]

In this study, the properties of circulating extracellular vesicles (EVs) were examined in cerebral palsy (CP) and typically developed (TD) individuals at rest and after aerobic exercise, focusing on the size, concentration, and microRNA cargo of EVs. Nine adult individuals with CP performed a single exercise bout consisting of 45 min of Frame Running, and TD participants completed either 45min of cycling (n = 10; TD EX) or were enrolled as controls with no exercise (n = 10; TD CON). Blood was drawn before and 30min after exercise and analyzed for EV concentration, size, and microRNA content. The size of EVs was similar in CP vs. TD, and exercise had no effect. Individuals with CP had an overall lower concentration (~25%, p < 0.05) of EVs. At baseline, let-7a, let-7b and let-7e were downregulated in individuals with CP compared to TD (p < 0.05), while miR-100 expression was higher, and miR-877 and miR-4433 lower in CP compared to TD after exercise (p < 0.05). Interestingly, miR-486 was upregulated ~2-fold in the EVs of CP vs. TD both at baseline and after exercise. We then performed an in silico analysis of miR-486 targets and identified the satellite cell stemness factor Pax7 as a target of miR-486. C2C12 myoblasts were cultured with amiR-486 mimetic and RNA-sequencing was performed. Gene enrichment analysis revealed that several genes involved in sarcomerogenesis and extracellular matrix (ECM) were downregulated. Our data suggest that circulating miR-486 transported by EVs is elevated in individuals with CP and that miR-486 alters the transcriptome of myoblasts affecting both ECM- and sarcomerogenesis-related genes, providing a link to the skeletal muscle alterations observed in individuals with CP. [ABSTRACT FROM AUTHOR]

In 1924, Otto Warburg asked "How does the metabolism of a growing tissue differ from that of a non-growing tissue?" Currently, we know that proliferating healthy and cancer cells reprogramme their metabolism. This typically includes increased glucose uptake, glycolytic flux and lactate synthesis. A key function of this reprogramming is to channel glycolytic intermediates and other metabolites into anabolic reactions such as nucleotide-RNA/DNA synthesis, amino acid-protein synthesis and the synthesis of, for example, acetyl and methyl groups for epigenetic modification. In this review, we discuss evidence that a hypertrophying muscle similarly takes up more glucose and reprogrammes its metabolism to channel energy metabolites into anabolic pathways. We specifically discuss the functions of the cancer-associated enzymes phosphoglycerate dehydrogenase and pyruvate kinase muscle 2 in skeletal muscle. In addition, we ask whether increased glucose uptake by a hypertrophying muscle explains why muscularity is often negatively associated with type 2 diabetes mellitus and obesity. [ABSTRACT FROM AUTHOR]

Many of the molecular and cellular mechanisms discovered to regulate skeletal muscle hypertrophy were first identified using the rodent synergist ablation model. This model reveals the intrinsic capability and necessary pathways of skeletal muscle growth in response to mechanical overload (MOV). Reminiscent of the rapid cellular growth observed with cancer, we hypothesized that in response to MOV, skeletal muscle would undergo metabolic programming to sustain increased demands to support hypertrophy. To test this hypothesis, we analyzed the gene expression of specific metabolic pathways taken from transcriptomic microarray data of a MOV time course. We found an upregulation of genes involved in the oxidative branch of the pentose phosphate pathways (PPP) and mitochondrial branch of the folate cycle suggesting an increase in the production of NADPH. In addition, we sought to determine the potential role of skeletal muscle‐enriched microRNA (myomiRs) and satellite cells in the regulation of the metabolic pathways that changed during MOV. We observed an inverse pattern in gene expression between muscle‐enriched myomiR‐1 and its known target gene glucose‐6‐phosphate dehydrogenase, G6pdx, suggesting myomiR regulation of PPP activation in response to MOV. Satellite cell fusion had a significant but modest impact on PPP gene expression. These transcriptomic findings suggest the robust muscle hypertrophy induced by MOV requires enhanced redox metabolism via PPP production of NADPH which is potentially regulated by a myomiR network. [ABSTRACT FROM AUTHOR]

There is emerging evidence of a gut microbiome–skeletal muscle axis. The purpose of this study was to determine if an intact gut microbiome was necessary for skeletal muscle adaptation to exercise. Forty‐two 4‐month‐old female C57BL/6J mice were randomly assigned to untreated (U) or antibiotic‐treated (T) non‐running controls (CU or CT, respectively) or progressive weighted wheel running (PoWeR, P) untreated (PU) or antibiotic‐treated (PT) groups. Antibiotic treatment resulted in disruption of the gut microbiome as indicated by a significant depletion of gut microbiome bacterial species in both CT and PT groups. The training stimulus was the same between PU and PT groups as assessed by weekly (12.35 ± 2.06 vs. 11.09 ± 1.76 km/week, respectively) and total (778.9 ± 130.5 vs. 703.8 ± 112.9 km, respectively) running activity. In response to PoWeR, PT showed less hypertrophy of soleus type 1 and 2a fibres and plantaris type 2b/x fibres compared to PU. The higher satellite cell and myonuclei abundance of PU plantaris muscle after PoWeR was not observed in PT. The fibre‐type shift of PU plantaris muscle to a more oxidative type 2a fibre composition following PoWeR was blunted in PT. There was no difference in serum cytokine levels among all groups suggesting disruption of the gut microbiome did not induce systemic inflammation. The results of this study provide the first evidence that an intact gut microbiome is necessary for skeletal muscle adaptation to exercise. Key points: Dysbiosis of the gut microbiome caused by continuous antibiotic treatment did not affect running activity.Continuous treatment with antibiotics did not result in systemic inflammation as indicated by serum cytokine levels.Gut microbiome dysbiosis was associated with blunted fibre type‐specific hypertrophy in the soleus and plantaris muscles in response to progressive weighted wheel running (PoWeR).Gut microbiome dysbiosis was associated with impaired PoWeR‐induced fibre‐type shift in the plantaris muscle.Gut microbiome dysbiosis was associated with a loss of PoWeR‐induced myonuclei accretion in the plantaris muscle. [ABSTRACT FROM AUTHOR]

Skeletal muscle in individuals with CP also contains lower amounts of mtDNA, potentially indicating fewer mitochondria in CP skeletal muscle compared with typically developing muscle. We compared skeletal muscle samples from children with cerebral palsy (CP) and typically developing children and observed evidence of reduced mtDNA and OXPHOS protein content in CP skeletal muscle, indicating reduced mitochondrial abundance. Cerebral palsy (CP) muscle contains fewer energy-generating organelles than typically developing muscle. [Extracted from the article]

Key points: Ribosome biogenesis and MYC transcription are associated with acute resistance exercise (RE) and are distinct from endurance exercise in human skeletal muscle throughout a 24 h time course of recovery.A PCR‐based method for relative ribosomal DNA (rDNA) copy number estimation was validated by whole genome sequencing and revealed that rDNA dosage is positively correlated with ribosome biogenesis in response to RE.Acute RE modifies rDNA methylation patterns in enhancer, intergenic spacer and non‐canonical MYC‐associated regions, but not the promoter.Myonuclear‐specific rDNA methylation patterns with acute mechanical overload in mice corroborate and expand on rDNA findings with RE in humans.A genetic predisposition for hypertrophic responsiveness may exist based on rDNA gene dosage. Ribosomes are the macromolecular engines of protein synthesis. Skeletal muscle ribosome biogenesis is stimulated by exercise, although the contribution of ribosomal DNA (rDNA) copy number and methylation to exercise‐induced rDNA transcription is unclear. To investigate the genetic and epigenetic regulation of ribosome biogenesis with exercise, a time course of skeletal muscle biopsies was obtained from 30 participants (18 men and 12 women; 31 ± 8 years, 25 ± 4 kg m–2) at rest and 30 min, 3 h, 8 h and 24 h after acute endurance (n = 10, 45 min cycling, 70% V̇O2max) or resistance exercise (n = 10, 4 × 7 × 2 exercises); 10 control participants underwent biopsies without exercise. rDNA transcription and dosage were assessed using quantitative PCR and whole genome sequencing. rDNA promoter methylation was investigated using massARRAY EpiTYPER and global rDNA CpG methylation was assessed using reduced‐representation bisulphite sequencing. Ribosome biogenesis and MYC transcription were associated primarily with resistance but not endurance exercise, indicating preferential up‐regulation during hypertrophic processes. With resistance exercise, ribosome biogenesis was associated with rDNA gene dosage, as well as epigenetic changes in enhancer and non‐canonical MYC‐associated areas in rDNA, but not the promoter. A mouse model of in vivo metabolic RNA labelling and genetic myonuclear fluorescence labelling validated the effects of an acute hypertrophic stimulus on ribosome biogenesis and Myc transcription, and also corroborated rDNA enhancer and Myc‐associated methylation alterations specifically in myonuclei. The present study provides the first information on skeletal muscle genetic and rDNA gene‐wide epigenetic regulation of ribosome biogenesis in response to exercise, revealing novel roles for rDNA dosage and CpG methylation. Key points: Ribosome biogenesis and MYC transcription are associated with acute resistance exercise (RE) and are distinct from endurance exercise in human skeletal muscle throughout a 24 h time course of recovery.A PCR‐based method for relative ribosomal DNA (rDNA) copy number estimation was validated by whole genome sequencing and revealed that rDNA dosage is positively correlated with ribosome biogenesis in response to RE.Acute RE modifies rDNA methylation patterns in enhancer, intergenic spacer and non‐canonical MYC‐associated regions, but not the promoter.Myonuclear‐specific rDNA methylation patterns with acute mechanical overload in mice corroborate and expand on rDNA findings with RE in humans.A genetic predisposition for hypertrophic responsiveness may exist based on rDNA gene dosage. [ABSTRACT FROM AUTHOR]

Regular exercise has a central role in human health by reducing the risk of type 2 diabetes, obesity, stroke and cancer. How exercise is able to promote such systemic benefits has remained somewhat of a mystery but has been thought to be in part mediated by the release of myokines, skeletal muscle‐specific cytokines, in response to exercise. Recent studies have revealed skeletal muscle can also release extracellular vesicles (EVs) into circulation following a bout of exercise. EVs are small membrane‐bound vesicles capable of delivering biomolecules to recipient cells and subsequently altering their metabolism. The notion that EVs may have a role in both skeletal muscle and systemic adaptation to exercise has generated a great deal of excitement within a number of different fields including exercise physiology, neuroscience and metabolism. The purpose of this review is to provide an introduction to EV biology and what is currently known about skeletal muscle EVs and their potential role in the response of muscle and other tissues to exercise. [ABSTRACT FROM AUTHOR]

Small noncoding microRNAs (miRNAs) are important regulators of skeletal muscle size, and circulating miRNAs within extracellular vesicles (EVs) may contribute to atrophy and its associated systemic effects. The purpose of this study was to understand how muscle atrophy and regrowth alter in vivo serum EV miRNA content. We also associated changes in serum EV miRNA with protein synthesis, protein degradation, and miRNA within muscle, kidney, and liver. We subjected adult (10 mo) F344/BN rats to three conditions: weight bearing (WB), hindlimb suspension (HS) for 7 days to induce muscle atrophy, and HS for 7 days followed by 7 days of reloading (HSR). Microarray analysis of EV miRNA content showed that the overall changes in serum EV miRNA were predicted to target major anabolic, catabolic, and mechanosensitive pathways. MiR-203a-3p was the only miRNA demonstrating substantial differences in HS EVs compared with WB. There was a limited association of EV miRNA content to the corresponding miRNA content within the muscle, kidney, or liver. Stepwise linear regression demonstrated that EV miR-203a-3p was correlated with muscle mass and muscle protein synthesis and degradation across all conditions. Finally, EV miR-203a-3p expression was significantly decreased in human subjects who underwent unilateral lower limb suspension (ULLS) to induce muscle atrophy. Altogether, we show that serum EV miR-203a-3p expression is related to skeletal muscle protein turnover and atrophy. We suggest that serum EV miR-203a-3p content may be a useful biomarker and future work should investigate whether serum EV miR-203a-3p content is mechanistically linked to protein synthesis and degradation. [ABSTRACT FROM AUTHOR]

To date, studies that have aimed to investigate the role of satellite cells during adult skeletal muscle adaptation and hypertrophy have utilized a nontranslational stimulus and/or have been performed over a relatively short time frame. Although it has been shown that satellite cell depletion throughout adulthood does not drive skeletal muscle loss in sedentary mice, it remains unknown how satellite cells participate in skeletal muscle adaptation to long-term physical activity. The current study was designed to determine whether reduced satellite cell content throughout adulthood would influence the transcriptome-wide response to physical activity and diminish the adaptive response of skeletal muscle. We administered vehicle or tamoxifen to adult Pax7-diphtheria toxin A (DTA) mice to deplete satellite cells and assigned them to sedentary or wheel-running conditions for 13 mo. Satellite cell depletion throughout adulthood reduced balance and coordination, overall running volume, and the size of muscle proprioceptors (spindle fibers). Furthermore, satellite cell participation was necessary for optimal muscle fiber hypertrophy but not adaptations in fiber type distribution in response to lifelong physical activity. Transcriptome-wide analysis of the plantaris and soleus revealed that satellite cell function is muscle type specific; satellite cell-dependent myonuclear accretion was apparent in oxidative muscles, whereas initiation of G protein-coupled receptor (GPCR) signaling in the glycolytic plantaris may require satellite cells to induce optimal adaptations to long-term physical activity. These findings suggest that satellite cells play a role in preserving physical function during aging and influence muscle adaptation during sustained periods of physical activity. [ABSTRACT FROM AUTHOR]

The current study investigated how bovine milk extracellular vesicles (EVs) affected rotarod performance and biomarkers of skeletal muscle physiology in young, growing rats. Twenty-eight-day Fisher 344 rats were provided an AIN-93G-based diet for 4 weeks that either remained unadulterated [EVs and RNA-sufficient (ERS; n = 12)] or was sonicated [EVs and RNA-depleted (ERD; n = 12)]. Prior to (PRE) and on the last day of the intervention (POST), animals were tested for maximal rotarod performance. Following the feeding period, the gastrocnemius muscle was analyzed at the histological, biochemical, and molecular levels and was also used to measure mitochondrial function and reactive oxygen species (ROS) emission. A main effect of time was observed for rotarod time (PRE > POST, p = 0.001). Terminal gastrocnemius mass was unaffected by diet, although gastrocnemius muscle fiber cross sectional area was 11% greater (p = 0.018) and total RNA (a surrogate of ribosome density) was 24% greater (p = 0.001) in ERD. Transcriptomic analysis of the gastrocnemius indicated that 22 mRNAs were significantly greater in ERS versus ERD (p < 0.01), whereas 55 mRNAs were greater in ERD versus ERS (p < 0.01). There were no differences in gastrocnemius citrate synthase activity or mitochondrial coupling (respiratory control ratio), although mitochondrial ROS production was lower in ERD gastrocnemius (p = 0.016), which may be explained by an increase in glutathione peroxidase protein levels (p = 0.020) in ERD gastrocnemius. Dietary EVs profiling confirmed that sonication in the ERD diet reduced EVs content by ∼60%. Our findings demonstrate that bovine milk EVs depletion through sonication elicits anabolic and transcriptomic effects in the gastrocnemius muscle of rapidly maturing rats. While this did not translate into a functional outcome between diets (i.e., rotarod performance), longer feeding periods may be needed to observe such functional effects. [ABSTRACT FROM AUTHOR]

We sought to characterize the lipid profile of skeletal muscle cell-derived Extracellular Vesicles (EVs) to determine if a hypertrophic stimulus would affect the lipid composition of C2C12 myotube-derived EVs. Analyses included C2C12 murine myoblasts differentiated into myotubes and treated with Insulin-Like Growth Factor 1 (IGF-1) for 24 h to induce hypertrophic growth. EVs were isolated from cell culture media, quantified using Nanoparticle Tracking Analysis (NTA) and analyzed using Transmission Electron Microscopy (TEM). EVs were homogenized and lipids extracted for quantification by Mass Spectrometry followed by downstream lipid class enrichment and lipid chain analysis. IGF-1 treatment elicited an increase in CD63 and CD81 levels (39% and 21%) compared to the controls (16%), respectively. Analysis revealed that skeletal muscle-derived EVs are enriched in bioactive lipids that are likely selectively incorporated into EVs during hypertrophic growth. IGF-1 treatment of myotubes had a significant impact on the levels of diacylglycerol (DG) and ceramide (Cer) in secreted EVs. Specifically, the proportion of unsaturated DG was two- to three-fold higher in EVs derived from IGF-treated cells, as compared to those from control cells. The levels of saturated DG were unaffected. Selective increases were similarly seen in C16- and C24-Cer but not in other species. Levels of free sphingoid bases tended to decrease, while those of sphingosine-1-phosphate was unaffected. Our results suggest that the lipid composition and biogenesis of skeletal muscle-derived EVs, are specific and highly selective during hypertrophic growth. [ABSTRACT FROM AUTHOR]

We generated an inducible, skeletal muscle-specific Dicer knockout mouse to deplete microRNAs in adult skeletal muscle. Following tamoxifen treatment, Dicer mRNA expression was significantly decreased by 87%. Wild-type (WT) and Dicer knockout (KO) mice were subjected to either synergist ablation or hind limb suspension for two weeks. There was no difference in muscle weight with hypertrophy or atrophy between WT and KO groups; however, even with the significant loss of Dicer expression, myomiR (miR-1, -133a and -206) expression was only reduced by 38% on average. We next aged WT and KO mice for ~22 months following Dicer inactivation to determine if myomiR expression would be further reduced over a prolonged timeframe and assess the effects of myomiR depletion on skeletal muscle phenotype. Skeletal muscle Dicer mRNA expression remained significantly decreased by 80% in old KO mice and sequencing of cloned Dicer mRNA revealed the complete absence of the floxed exons in KO skeletal muscle. Despite a further reduction of myomiR expression to ~50% of WT, no change was observed in muscle morphology between WT and KO groups. These results indicate the life-long reduction in myomiR levels did not adversely affect skeletal muscle phenotype and suggest the possibility that microRNA expression is uniquely regulated in skeletal muscle. [ABSTRACT FROM AUTHOR]

Background: The tetracycline-responsive system (Tet-ON/OFF) has proven to be a valuable tool for manipulating gene expression in an inducible, temporal, and tissue-specific manner. The purpose of this study was to create and characterize a new transgenic mouse strain utilizing the human skeletal muscle α-actin (HSA) promoter to drive skeletal muscle-specific expression of the reverse tetracycline transactivator (rtTA) gene which we have designated as the HSA-rtTA mouse. Methods: To confirm the HSA-rtTA mouse was capable of driving skeletal muscle-specific expression, we crossed the HSA-rtTA mouse with the tetracycline-responsive histone H2B-green fluorescent protein (H2B-GFP) transgenic mouse in order to label myonuclei. Results: Reverse transcription-PCR confirmed skeletal muscle-specific expression of rtTA mRNA, while single-fiber analysis showed highly effective GFP labeling of myonuclei in both fast- and slow-twitch skeletal muscles. Pax7 immunohistochemistry of skeletal muscle cross-sections revealed no appreciable GFP expression in satellite cells. Conclusions: The HSA-rtTA transgenic mouse allows for robust, specific, and inducible gene expression across muscles of different fiber types. The HSA-rtTA mouse provides a powerful tool to manipulate gene expression in skeletal muscle. [ABSTRACT FROM AUTHOR]