Am J Physiol Cell Physiol 307: C311–C313, 2014; doi:10.1152/ajpcell.00219.2014.
Editorial Focus
Out FoxO’d by microRNA. Focus on “miR-182 attenuates atrophy-related gene expression by targeting FoxO3 in skeletal muscle” John J. McCarthy Department of Physiology and Center for Muscle Biology, University of Kentucky, Lexington, Kentucky OVER THE PAST FEW YEARS, non-coding RNAs (ncRNAs) have emerged as important regulators of gene expression (2). A family of small ncRNAs, known as microRNAs (miRNAs), have been shown to have a role in a wide range of developmental and cellular processes as well as certain pathologies such as cancer. This family of ncRNAs functions through a posttranscriptional mechanism by inhibiting mRNA translation and/or degradation of a target transcript. The discovery that miRNAs were present in a wide range of animals also revealed that some miRNAs were in fact expressed in a tissue-specific manner; for example, the expression of the mature miRNA (miR) miR-1 was found to be restricted to the heart, with later studies showing miR-1 expression in skeletal muscle as well (9). Since then, a small family of muscle-specific miRNAs, referred to as myomiRs, have been identified and shown to have a central role in myogenesis and adaptation to both physiological and pathological stresses (7).
Address for reprint requests and other correspondence: J. J. McCarthy, Dept. of Physiology, Univ. of Kentucky, 800 Rose St., Lexington, KY 40536-0298 (e-mail: [email protected]). IGF-1 IGFR-1
myostatin
The finding that miRNAs might mediate skeletal muscle plasticity is not surprising given the general consensus in the field that miRNAs are involved in mediating the stress response of the cell, helping to maintain and restore cellular homeostasis through the regulation of gene expression (3). In recent years, evidence has emerged that both myomiRs and more broadly expressed miRNAs are involved in the regulation of skeletal muscle metabolism, fiber type, and mass (7). In particular, great strides have been made in understanding how miRNAs impact the regulatory networks that underlie skeletal muscle atrophy in response to catabolic stimuli. The muscle-specific E3 ubiquitin ligases, MAFbx/atrogin-1 and muscle RING-finger 1 (MuRF-1), are master regulators of skeletal muscle atrophy by increasing cellular protein degradation via targeting proteins for the ubiquitin-proteasome system (5). The finding that the forkhead box O3 (FoxO3) transcription factor promotes expression of MAFbx/atrogin-1 and MuRF-1, as well as genes involved in autophagic/lysosomal proteolysis, provided a unifying mechanism for coordinating the regulation of protein degradation that underlies muscle atrophy (10, 15). Wada and colleagues (14) provided the first miR-27a/b
ActIIRβ
PI3K
AKT
HSP70
FoxO3
MAFbx
miR-1
miR-182
Fig. 1. MicroRNA regulation of skeletal muscle atrophy. Schematic of some of the miRNAs reported to target genes involved in skeletal muscle atrophy: miR27a/b was shown to repress expression of myostatin; the muscle-specific E3 ubiquitin ligases MAFbx/atrogin-1 and MuRF-1 have been validated as target genes of miR-23a; the muscle-specific miR-1 targets HSP70 thereby affecting Akt activity; the current study provides evidence that miR-182 directly targets FoxO3 in skeletal muscle.
MuRF-1
miR-23a protein degradation
muscle atrophy http://www.ajpcell.org
0363-6143/14 Copyright © 2014 the American Physiological Society
C311
Editorial Focus C312 evidence that a miRNA could affect this signaling pathway by inhibiting the translation of both MAFbx/atrogin-1 and MuRF-1 mRNAs (Fig. 1). These authors showed that miR-23a was capable of repressing expression of MAFbx/atrogin-1 and MuRF-1 and that miR-23a overexpression protected mice against glucocorticoid-induced muscle atrophy (14). In an earlier study, Allen and Loh (1) demonstrated that the increase in myostatin protein following glucocorticoid treatment was caused by a downregulation of miR-27a/b expression. Though it remains to be fully explored, together these findings suggest that miR-23a and miR-27a/b have a central role in skeletal muscle atrophy through their regulation of these atrogenes. More recently, Kukreti and colleagues (8) reported that muscle-specific miR-1 was involved in mediating dexamethasone-induced muscle atrophy by enhancing FoxO3 activity, ultimately resulting in the upregulation of MAFbx/atrogin-1 and MuRF-1 expression. They showed that glucocorticoid receptor activation of miR-1 expression resulted in decreased expression of the target gene HSP70, which, in turn, led to FoxO3 activation via reduced Akt phosphorylation (8). Consistent with these findings, Senf et al. (12) found that HSP70 was able to directly block FoxO3 activation of the MAFbx/ atrogin-1 promoter reporter gene. The current study by Hudson and colleagues (6) extends these earlier findings in an important way by identifying a miRNA that directly targets FoxO3 (6). The authors focused on miR-182 based on the presence of two predicted miR-182 target sequences in the 3=-UTR of FoxO3 and a previous study that validated FoxO3 as a bona fide target of miR-182 in a melanoma cell line (11). Given the recent finding revealing the widespread context dependency of miRNA-mediated regulation, it was critical that the authors showed that in fact miR-182 was capable of repressing FoxO3 expression in muscle cells (4). Moreover, the expected increase in FoxO3 expression in myotubes following exposure to dexamethasone was associated with a significant downregulation of miR-182 expression. As further proof of the inverse relationship between FoxO3 and miR-182 expression, the authors employed an animal model of muscle atrophy associated with diabetes; the 75% increase in FoxO3 mRNA in diabetic muscle was associated with a 45% decrease in miR-182 expression, thus confirming the in vitro findings. The finding that miR-182 can repress FoxO3 expression suggests that the overexpression of miR-182 may provide a means to ameliorate glucocorticoid-induced muscle atrophy by preventing the activation of MAFbx/atrogin-1 and MuRF-1 by FoxO3. To test this exciting possibility, the authors showed that overexpression of miR-182 was able to prevent the increase in MAFbx/atrogin-1 expression in response to dexamethasone. Consistent with miR-182 regulation of FoxO3 expression, the authors also showed that the induction of other FoxO3 target genes involved in the autophagy/lysosome system was blocked by the overexpression of miR182. Together the findings of the study by Hudson and colleagues provide the foundation for exploring the therapeutic potential of miR-182 to prevent skeletal muscle atrophy under catabolic conditions (6). It will be important for future studies to determine in vivo just how effective the manipulation of miR-182 expression is for treating skeletal muscle atrophy under different conditions. Though the authors expressed some concern that
miR-182 was not as abundant as some myomiRs, calling into question the biological relevance of their findings, they have subsequently found by RNA-seq analysis that miR-182 is significantly more abundant than previously reported (personal communication, S.R.P.). Of some concern is the recent study by Soares and coworkers (13) that reported the miRNA expression profile associated with different skeletal muscle catabolic conditions (i.e., fasting, denervation, cachexia, and diabetes) was unique and that there did not appear to be a miRNA signature of muscle atrophy. Despite this caveat, given the central role that FoxO3 has in the regulation of skeletal muscle atrophy, the current study’s findings provide optimism for continued exploration of the use of miR-182 to treat muscle atrophy. DISCLOSURES No conflicts of interest, financial or otherwise, are declared by the author. AUTHOR CONTRIBUTIONS J.J.M. prepared figure; drafted manuscript; approved final version of manuscript. REFERENCES 1. Allen DL, Loh AS. Posttranscriptional mechanisms involving microRNA-27a and b contribute to fast-specific and glucocorticoid-mediated myostatin expression in skeletal muscle. Am J Physiol Cell Physiol 300: C124 –C137, 2010. 2. Cech TR, Steitz JA. The noncoding RNA revolution-trashing old rules to forge new ones. Cell 157: 77–94, 2014. 3. Ebert MS, Sharp PA. Roles for microRNAs in conferring robustness to biological processes. Cell 149: 515–524, 2012. 4. Erhard F, Haas J, Lieber D, Malterer G, Jaskiewicz L, Zavolan M, Dolken L, Zimmer R. Widespread context dependency of microRNAmediated regulation. Genome Res 24: 906 –919, 2014. 5. Foletta VC, White LJ, Larsen AE, Leger B, Russell AP. The role and regulation of MAFbx/atrogin-1 and MuRF1 in skeletal muscle atrophy. Pflügers Arch 461: 325–335, 2011. 6. Hudson MB, Rahnert JA, Zheng B, Woodworth-Hobbs ME, Franch HA, Price SR. miR-182 attenuates atrophy-related gene expression by targeting FoxO3 in skeletal muscle. Am J Physiol Cell Physiol (May 28, 2014). doi: 10.1152/ajpcell.00395.2013. 7. Kirby TJ, McCarthy JJ. MicroRNAs in skeletal muscle biology and exercise adaptation. Free Radic Biol Med 64: 95–105, 2013. 8. Kukreti H, Amuthavalli K, Harikumar A, Sathiyamoorthy S, Feng PZ, Anantharaj R, Tan SL, Lokireddy S, Bonala S, Sriram S, McFarlane C, Kambadur R, Sharma M. Muscle-specific microRNA1 (miR1) targets heat shock protein 70 (HSP70) during dexamethasonemediated atrophy. J Biol Chem 288: 6663–6678, 2013. 9. Lagos-Quintana M, Rauhut R, Yalcin A, Meyer J, Lendeckel W, Tuschl T. Identification of tissue-specific microRNAs from mouse. Curr Biol 12: 735–739, 2002. 10. Sandri M, Sandri C, Gilbert A, Skurk C, Calabria E, Picard A, Walsh K, Schiaffino S, Lecker SH, Goldberg AL. Foxo transcription factors induce the atrophy-related ubiquitin ligase atrogin-1 and cause skeletal muscle atrophy. Cell 117: 399 –412, 2004. 11. Segura MF, Hanniford D, Menendez S, Reavie L, Zou X, AlvarezDiaz S, Zakrzewski J, Blochin E, Rose A, Bogunovic D, Polsky D, Wei J, Lee P, Belitskaya-Levy I, Bhardwaj N, Osman I, Hernando E. Aberrant miR-182 expression promotes melanoma metastasis by repressing FOXO3 and microphthalmia-associated transcription factor. Proc Natl Acad Sci USA 106: 1814 –1819, 2009. 12. Senf SM, Dodd SL, Judge AR. FOXO signaling is required for disuse muscle atrophy and is directly regulated by Hsp70. Am J Physiol Cell Physiol 298: C38 –C45, 2010.
AJP-Cell Physiol • doi:10.1152/ajpcell.00219.2014 • www.ajpcell.org
Editorial Focus C313 13. Soares RJ, Cagnin S, Chemello F, Silvestrin M, Musaro A, De Pitta C, Lanfranchi G, Sandri M. Involvement of miRNAs in the regulation of muscle wasting during catabolic conditions. J Biol Chem doi: 10.1074/ jbc.M114.561845 [Epub ahead of print]. 14. Wada S, Kato Y, Okutsu M, Miyaki S, Suzuki K, Yan Z, Schiaffino S, Asahara H, Ushida T, Akimoto T. Translational suppression of atrophic
regulators by microRNA-23a integrates resistance to skeletal muscle atrophy. J Biol Chem 286: 38456 –38465, 2011. 15. Zhao J, Brault JJ, Schild A, Cao P, Sandri M, Schiaffino S, Lecker SH, Goldberg AL. FoxO3 coordinately activates protein degradation by the autophagic/lysosomal and proteasomal pathways in atrophying muscle cells. Cell Metab 6: 472–483, 2007.
AJP-Cell Physiol • doi:10.1152/ajpcell.00219.2014 • www.ajpcell.org
Copyright of American Journal of Physiology: Cell Physiology is the property of American Physiological Society and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
Editorial Focus
Out FoxO’d by microRNA. Focus on “miR-182 attenuates atrophy-related gene expression by targeting FoxO3 in skeletal muscle” John J. McCarthy Department of Physiology and Center for Muscle Biology, University of Kentucky, Lexington, Kentucky OVER THE PAST FEW YEARS, non-coding RNAs (ncRNAs) have emerged as important regulators of gene expression (2). A family of small ncRNAs, known as microRNAs (miRNAs), have been shown to have a role in a wide range of developmental and cellular processes as well as certain pathologies such as cancer. This family of ncRNAs functions through a posttranscriptional mechanism by inhibiting mRNA translation and/or degradation of a target transcript. The discovery that miRNAs were present in a wide range of animals also revealed that some miRNAs were in fact expressed in a tissue-specific manner; for example, the expression of the mature miRNA (miR) miR-1 was found to be restricted to the heart, with later studies showing miR-1 expression in skeletal muscle as well (9). Since then, a small family of muscle-specific miRNAs, referred to as myomiRs, have been identified and shown to have a central role in myogenesis and adaptation to both physiological and pathological stresses (7).
Address for reprint requests and other correspondence: J. J. McCarthy, Dept. of Physiology, Univ. of Kentucky, 800 Rose St., Lexington, KY 40536-0298 (e-mail: [email protected]). IGF-1 IGFR-1
myostatin
The finding that miRNAs might mediate skeletal muscle plasticity is not surprising given the general consensus in the field that miRNAs are involved in mediating the stress response of the cell, helping to maintain and restore cellular homeostasis through the regulation of gene expression (3). In recent years, evidence has emerged that both myomiRs and more broadly expressed miRNAs are involved in the regulation of skeletal muscle metabolism, fiber type, and mass (7). In particular, great strides have been made in understanding how miRNAs impact the regulatory networks that underlie skeletal muscle atrophy in response to catabolic stimuli. The muscle-specific E3 ubiquitin ligases, MAFbx/atrogin-1 and muscle RING-finger 1 (MuRF-1), are master regulators of skeletal muscle atrophy by increasing cellular protein degradation via targeting proteins for the ubiquitin-proteasome system (5). The finding that the forkhead box O3 (FoxO3) transcription factor promotes expression of MAFbx/atrogin-1 and MuRF-1, as well as genes involved in autophagic/lysosomal proteolysis, provided a unifying mechanism for coordinating the regulation of protein degradation that underlies muscle atrophy (10, 15). Wada and colleagues (14) provided the first miR-27a/b
ActIIRβ
PI3K
AKT
HSP70
FoxO3
MAFbx
miR-1
miR-182
Fig. 1. MicroRNA regulation of skeletal muscle atrophy. Schematic of some of the miRNAs reported to target genes involved in skeletal muscle atrophy: miR27a/b was shown to repress expression of myostatin; the muscle-specific E3 ubiquitin ligases MAFbx/atrogin-1 and MuRF-1 have been validated as target genes of miR-23a; the muscle-specific miR-1 targets HSP70 thereby affecting Akt activity; the current study provides evidence that miR-182 directly targets FoxO3 in skeletal muscle.
MuRF-1
miR-23a protein degradation
muscle atrophy http://www.ajpcell.org
0363-6143/14 Copyright © 2014 the American Physiological Society
C311
Editorial Focus C312 evidence that a miRNA could affect this signaling pathway by inhibiting the translation of both MAFbx/atrogin-1 and MuRF-1 mRNAs (Fig. 1). These authors showed that miR-23a was capable of repressing expression of MAFbx/atrogin-1 and MuRF-1 and that miR-23a overexpression protected mice against glucocorticoid-induced muscle atrophy (14). In an earlier study, Allen and Loh (1) demonstrated that the increase in myostatin protein following glucocorticoid treatment was caused by a downregulation of miR-27a/b expression. Though it remains to be fully explored, together these findings suggest that miR-23a and miR-27a/b have a central role in skeletal muscle atrophy through their regulation of these atrogenes. More recently, Kukreti and colleagues (8) reported that muscle-specific miR-1 was involved in mediating dexamethasone-induced muscle atrophy by enhancing FoxO3 activity, ultimately resulting in the upregulation of MAFbx/atrogin-1 and MuRF-1 expression. They showed that glucocorticoid receptor activation of miR-1 expression resulted in decreased expression of the target gene HSP70, which, in turn, led to FoxO3 activation via reduced Akt phosphorylation (8). Consistent with these findings, Senf et al. (12) found that HSP70 was able to directly block FoxO3 activation of the MAFbx/ atrogin-1 promoter reporter gene. The current study by Hudson and colleagues (6) extends these earlier findings in an important way by identifying a miRNA that directly targets FoxO3 (6). The authors focused on miR-182 based on the presence of two predicted miR-182 target sequences in the 3=-UTR of FoxO3 and a previous study that validated FoxO3 as a bona fide target of miR-182 in a melanoma cell line (11). Given the recent finding revealing the widespread context dependency of miRNA-mediated regulation, it was critical that the authors showed that in fact miR-182 was capable of repressing FoxO3 expression in muscle cells (4). Moreover, the expected increase in FoxO3 expression in myotubes following exposure to dexamethasone was associated with a significant downregulation of miR-182 expression. As further proof of the inverse relationship between FoxO3 and miR-182 expression, the authors employed an animal model of muscle atrophy associated with diabetes; the 75% increase in FoxO3 mRNA in diabetic muscle was associated with a 45% decrease in miR-182 expression, thus confirming the in vitro findings. The finding that miR-182 can repress FoxO3 expression suggests that the overexpression of miR-182 may provide a means to ameliorate glucocorticoid-induced muscle atrophy by preventing the activation of MAFbx/atrogin-1 and MuRF-1 by FoxO3. To test this exciting possibility, the authors showed that overexpression of miR-182 was able to prevent the increase in MAFbx/atrogin-1 expression in response to dexamethasone. Consistent with miR-182 regulation of FoxO3 expression, the authors also showed that the induction of other FoxO3 target genes involved in the autophagy/lysosome system was blocked by the overexpression of miR182. Together the findings of the study by Hudson and colleagues provide the foundation for exploring the therapeutic potential of miR-182 to prevent skeletal muscle atrophy under catabolic conditions (6). It will be important for future studies to determine in vivo just how effective the manipulation of miR-182 expression is for treating skeletal muscle atrophy under different conditions. Though the authors expressed some concern that
miR-182 was not as abundant as some myomiRs, calling into question the biological relevance of their findings, they have subsequently found by RNA-seq analysis that miR-182 is significantly more abundant than previously reported (personal communication, S.R.P.). Of some concern is the recent study by Soares and coworkers (13) that reported the miRNA expression profile associated with different skeletal muscle catabolic conditions (i.e., fasting, denervation, cachexia, and diabetes) was unique and that there did not appear to be a miRNA signature of muscle atrophy. Despite this caveat, given the central role that FoxO3 has in the regulation of skeletal muscle atrophy, the current study’s findings provide optimism for continued exploration of the use of miR-182 to treat muscle atrophy. DISCLOSURES No conflicts of interest, financial or otherwise, are declared by the author. AUTHOR CONTRIBUTIONS J.J.M. prepared figure; drafted manuscript; approved final version of manuscript. REFERENCES 1. Allen DL, Loh AS. Posttranscriptional mechanisms involving microRNA-27a and b contribute to fast-specific and glucocorticoid-mediated myostatin expression in skeletal muscle. Am J Physiol Cell Physiol 300: C124 –C137, 2010. 2. Cech TR, Steitz JA. The noncoding RNA revolution-trashing old rules to forge new ones. Cell 157: 77–94, 2014. 3. Ebert MS, Sharp PA. Roles for microRNAs in conferring robustness to biological processes. Cell 149: 515–524, 2012. 4. Erhard F, Haas J, Lieber D, Malterer G, Jaskiewicz L, Zavolan M, Dolken L, Zimmer R. Widespread context dependency of microRNAmediated regulation. Genome Res 24: 906 –919, 2014. 5. Foletta VC, White LJ, Larsen AE, Leger B, Russell AP. The role and regulation of MAFbx/atrogin-1 and MuRF1 in skeletal muscle atrophy. Pflügers Arch 461: 325–335, 2011. 6. Hudson MB, Rahnert JA, Zheng B, Woodworth-Hobbs ME, Franch HA, Price SR. miR-182 attenuates atrophy-related gene expression by targeting FoxO3 in skeletal muscle. Am J Physiol Cell Physiol (May 28, 2014). doi: 10.1152/ajpcell.00395.2013. 7. Kirby TJ, McCarthy JJ. MicroRNAs in skeletal muscle biology and exercise adaptation. Free Radic Biol Med 64: 95–105, 2013. 8. Kukreti H, Amuthavalli K, Harikumar A, Sathiyamoorthy S, Feng PZ, Anantharaj R, Tan SL, Lokireddy S, Bonala S, Sriram S, McFarlane C, Kambadur R, Sharma M. Muscle-specific microRNA1 (miR1) targets heat shock protein 70 (HSP70) during dexamethasonemediated atrophy. J Biol Chem 288: 6663–6678, 2013. 9. Lagos-Quintana M, Rauhut R, Yalcin A, Meyer J, Lendeckel W, Tuschl T. Identification of tissue-specific microRNAs from mouse. Curr Biol 12: 735–739, 2002. 10. Sandri M, Sandri C, Gilbert A, Skurk C, Calabria E, Picard A, Walsh K, Schiaffino S, Lecker SH, Goldberg AL. Foxo transcription factors induce the atrophy-related ubiquitin ligase atrogin-1 and cause skeletal muscle atrophy. Cell 117: 399 –412, 2004. 11. Segura MF, Hanniford D, Menendez S, Reavie L, Zou X, AlvarezDiaz S, Zakrzewski J, Blochin E, Rose A, Bogunovic D, Polsky D, Wei J, Lee P, Belitskaya-Levy I, Bhardwaj N, Osman I, Hernando E. Aberrant miR-182 expression promotes melanoma metastasis by repressing FOXO3 and microphthalmia-associated transcription factor. Proc Natl Acad Sci USA 106: 1814 –1819, 2009. 12. Senf SM, Dodd SL, Judge AR. FOXO signaling is required for disuse muscle atrophy and is directly regulated by Hsp70. Am J Physiol Cell Physiol 298: C38 –C45, 2010.
AJP-Cell Physiol • doi:10.1152/ajpcell.00219.2014 • www.ajpcell.org
Editorial Focus C313 13. Soares RJ, Cagnin S, Chemello F, Silvestrin M, Musaro A, De Pitta C, Lanfranchi G, Sandri M. Involvement of miRNAs in the regulation of muscle wasting during catabolic conditions. J Biol Chem doi: 10.1074/ jbc.M114.561845 [Epub ahead of print]. 14. Wada S, Kato Y, Okutsu M, Miyaki S, Suzuki K, Yan Z, Schiaffino S, Asahara H, Ushida T, Akimoto T. Translational suppression of atrophic
regulators by microRNA-23a integrates resistance to skeletal muscle atrophy. J Biol Chem 286: 38456 –38465, 2011. 15. Zhao J, Brault JJ, Schild A, Cao P, Sandri M, Schiaffino S, Lecker SH, Goldberg AL. FoxO3 coordinately activates protein degradation by the autophagic/lysosomal and proteasomal pathways in atrophying muscle cells. Cell Metab 6: 472–483, 2007.
AJP-Cell Physiol • doi:10.1152/ajpcell.00219.2014 • www.ajpcell.org
Copyright of American Journal of Physiology: Cell Physiology is the property of American Physiological Society and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
