Overview
Molecular Mechanisms that Impact Transcription of Myosin Genes in Human Skeletal Muscle in Response to Unloading: Role of Resistance Exercise (Postdoctoral Fellowship)
Principal Investigator:
Clay E. Pandorf, Ph.D.
Organization:
University of California, Irvine
The reduction of muscle mass is a health concern for astronauts during a long-duration spaceflight. Muscle fiber properties and endurance capacities also undergo changes in the reduced-gravity environment of space. The alterations to muscle fibers are attributed to modification of myosin heavy chain (MHC) genes, which are the major contractile proteins found in muscle fibers.
Dr. Clay Pandorf’s project will examine the mechanisms that cause the changes in muscle genes in humans during simulated spaceflight conditions. The project will also look at the effectiveness of using a flywheel resistance exercise device in reducing the muscle fiber changes. The results of this study could help develop future countermeasures that will reduce muscle loss during spaceflight, as well as muscle loss on Earth due to disuse.
NASA Taskbook Entry
Technical Summary
Loading forces are critical to maintaining homeostasis in the muscle cell. Perturbations, such as microgravity, upset the normal gene expression of structural and signaling proteins. Unloading of skeletal muscle results in atrophy as well as deleterious alterations to the contractile properties and endurance capacities of muscle fibers. Exercise countermeasures can offset these alterations. Human skeletal muscle responds to exercise stimuli with alterations to myosin heavy chain (MHC) gene expression, and thus, contractile function.
Hypotheses
1. Does antisense RNA play a role in the transcriptional regulation of MHC genes in human skeletal muscle?
2. Are histone modifications dynamically regulated at MHC genes during alterations to muscle loading state in human skeletal muscle?
3. What are the functional performance and cellular-level adaptations to unloading and exercise training, and how does that relate to the corresponding molecular-level responses?
Key Findings
Vastus lateralis biopsies were obtained before and after five weeks of aerobic + resistance exercise (12 M, 9 F). The proportions of Type I and IIa MHC mRNA were both increased by 11%, while IIx was decreased by 38% after exercise training (all p < .05) in the vastus lateralis. Several novel discoveries were made regarding expression of long non-coding antisense RNA to MHC genes. As previously shown only in rats, lncRNA oriented antisense to the IIa MHC was detected and found to be down regulated with exercise (-55%; p < .05). A regulatory role is inferred from an inverse correlation between the proportion of IIa MHC mRNA and its antisense pair (r = -0.66; p = .002). Surprisingly, the researchers detected low levels of IIb mRNA and pre-mRNA, indicating that the IIb gene is not transcriptionally silent in human limb muscle, as thought. We also report the novel finding of as-lncRNA to the IIb gene. These data reveal that insights to MHC regulatory mechanisms previously shown in rats and involving as-lncRNA are conserved in humans.
Two groups of healthy inactive human subjects participated in unilateral lower-limb suspension (ULLS; male N=5; female N=4) for 10 days alone or with a combination of aerobic and resistance exercise training (ULLS+T; male N=5; female N=5). Soleus biopsies were obtained before and after ULLS. The researchers examined gene expression of several large sarcomeric proteins that provide structural support to the myofilament (titin, nebulin and α-actin) and converge at the Z-disc. They also examined several signaling proteins that reside in this region (STARS, atrogin-1, and calcineurin via its modulator MCIP1). Nebulin, titin and α-actin RNA levels changed pre to post by -11%, +26% and +7% with ULLS, and by +25%*, +56%* and +16% with ULLS+T, respectively (*p<0.05 pre to post). STARS, atrogin1 and MCIP1 mRNA levels changed pre to post by -75%*, +77%* and -40%* with ULLS, and by -63%*, +30%* and -26%* with ULLS+T, respectively (*p<0.05 pre to post). While changes in transcription of the Z-disc associated structural proteins nebulin and titin following10d unloading was not statistically significant, there was significant upregulation with unloading + training stimuli. The unfavorable unloading-induced signaling response was ameliorated when combined with training as suggested by differential mRNA levels of mechano-sensitive Z-disc factors that can promote atrophy (atrogin) and fiber-type shifts (calcineurin/MCIP1). These data suggest that exercise countermeasures to short-term unloading of the loading-sensitive slow soleus muscle can promote favorable transcriptional responses of proteins associated with the stress-sensitive Z-disc.
In conclusion, these studies showed that a gravity-independent exercise device can induce alterations to key contractile and structural muscle genes, and can counteract some of the early alterations that occur with simulated microgravity.
Earth Applications
Unloading of skeletal muscle results in atrophy as well as deleterious alterations to the contractile properties and endurance capacities of muscle fibers. This condition has significant impact on all humans with impaired muscle function due to disuse. These deleterious alterations are largely attributed to modification to the expression of the myosin heavy chain (MHC) genes. In particular non-fatigable, or slow, skeletal muscle motor units, which provide for postural support and sustained contractile activity, are remodeled to fatigable, or fast, muscle involving shifts from type I/IIa to IIx MHC in humans. Resistance exercise has been shown to effectively blunt these undesirable adaptations in both animals and humans.
The researchers have previously identified several novel mechanisms, such as antisense ribonucleic acid (RNA) and histone modifications that impact expression of the MHC genes in unloaded rat muscle. Specifically, the researchers showed that antisense RNA is implicated in transcriptional down-regulation of IIa MHC gene during muscle inactivity. They also demonstrated that alterations to histone acetylation and methylation at specific MHC genes correspond with altered MHC gene transcription in response to muscle unloading. The researchers also demonstrated that alterations to histone acetylation and methylation at specific MHC genes correspond with altered MHC gene transcription in response to muscle unloading.
In the present studies, the researchers were able to demonstrate that insights to MHC regulatory mechanisms previously shown in rats and involving long non-coding antisense RNA are conserved in humans. This may provide for development of novel therapeutics to curtail the slow to fast shift that occurs in disuse environments. They were also able to show that exercise countermeasures to short-term unloading of the loading-sensitive slow soleus muscle can promote favorable transcriptional responses of structural and signaling proteins in the muscle.
Validation with humans of the mechanisms that regulate the genes of contractile and structural proteins in muscle should help to develop future strategies that most effectively ameliorate the effects of unloading on human skeletal muscle, as well as enhancing the understanding of these same processes that have significant impact on all humans with impaired muscle function.
This project's funding ended in 2011
