MyoD and E-protein heterodimers switch rhabdomyosarcoma cells from an arrested myoblast phase to a differentiated state

Yang, Zhihong; MacQuarrie, Kyle L.; Analau, Erwin; Tyler, Ashlee E.; Dilworth, F. Jeffrey; Cao, Yi; Diede, Scott J.; Tapscott, Stephen J.

doi:10.1101/gad.1765109

Cited by 89 publications

(104 citation statements)

References 58 publications

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“…However in RMS, it is non-functional due to the formation of inhibitory dimers. 8,46 In addition, it has been shown that NF-kB and its target gene YY1 are highly expressed in RMS and epigenetically downregulate miR-29 b2/c. 15 In this study, we noticed that RMS cell lines showed variable responses to the ectopic expression of miR-1, -206 or -29.…”

Section: Discussionmentioning

confidence: 99%

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Downregulation of microRNAs miR-1, -206 and -29 stabilizes PAX3 and CCND2 expression in rhabdomyosarcoma

Sarver

Alamgir

et al. 2012

Laboratory Investigation

105

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Elevated levels of PAX3 and cell proliferation genes are characteristic features of rhabdomyosarcoma (RMS). We hypothesize that the increased levels of these genes are stabilized due to downregulation of specific miRNAs. In this study, we show that downregulation of miR-1, -206 and -29 stabilizes the expression of PAX3 and CCND2 in both embryonal (ERMS) and alveolar (ARMS) RMS types. Ectopic expression of miR-1 and 206 in JR1, an ERMS cell line, show significant downregulation of PAX3 protein expression, whereas overexpression of these miRNAs in Rh30, an ARMS cell line, did not show any effect in PAX3 protein levels. In ARMS, PAX3 forms a fusion transcript with FOXO1 and the resultant loss of PAX3 3 0 UTR in the fusion transcript indicate an oncogenic mechanism to evade miRNA-mediated regulation of PAX3. Further, we show that miR-1, -206 and -29 can regulate the expression of CCND2, a cell cycle gene. In addition to CCND2, miR-29 also targets E2F7, another cell cycle regulator. Cell function analysis shows that overexpression of miR-29 downregulates the expression of these cell cycle genes, induces partial G1 arrest leading to decreased cell proliferation. Taken together our data suggest that the RMS state is stabilized by the deregulation of multiple miRNAs and their target genes, supporting a tumor suppressor role for these miRNA. KEYWORDS: cell cycle genes; miRNAs; PAX3; regulatory networks; rhabdomyosarcoma Rhabdomyosarcoma (RMS) is a malignant striated muscle tumor that accounts for B3% of all childhood cancers. 1 RMS arises from primitive muscle cells and tumors show varying degrees of skeletal muscle differentiation that serves to define their classification as either embryonal (ERMS) or alveolar (ARMS) types. 2,3 ERMS, the most common type has features of embryonic muscle and are generally associated with favorable prognosis. In contrast, ARMS display poor muscle differentiation and are associated with poor outcomes. 4 Most ARMS cases are characterized by chromosomal translocation t(2;13) or t(1;13) involving genes PAX3-FKHR or PAX7-FKHR, respectively. 5 Regulatory disruptions in growth and differentiation pathways of myogenic precursor cells have been implicated in RMS development. Genes involved with muscle cell differentiation and cell proliferation have been associated with RMS development and metastasis. 6-11 Gene expression profiles comparing PAX-FOXO1-positive ARMS vs translocation-negative ERMS have identified genes relevant to tumorigenic process of ARMS and ERMS. 12 microRNAs (miRNAs) have been implicated in RMS. [13][14][15] miRNAs are small (18-22 nucleotides) evolutionarily conserved, non-coding RNAs that post-transcriptionally regulate gene expression through mRNA degradation, translation inhibition or chromatin-based silencing mechanisms. 16 Each miRNA can potentially regulate hundreds of targets either directly or indirectly. miRNAs have been shown to be deregulated in many types of cancers. 17 As a consequence, miRNAs from tumor tissue have been proposed for use in the diagnosis, classif...

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Section: Discussionmentioning

confidence: 99%

“…Genes involved with muscle cell differentiation and cell proliferation have been associated with RMS development and metastasis. [6][7][8][9][10][11] Gene expression profiles comparing PAX-FOXO1-positive ARMS vs translocation-negative ERMS have identified genes relevant to tumorigenic process of ARMS and ERMS. 12 microRNAs (miRNAs) have been implicated in RMS.…”

mentioning

confidence: 99%

Downregulation of microRNAs miR-1, -206 and -29 stabilizes PAX3 and CCND2 expression in rhabdomyosarcoma

Sarver

Alamgir

et al. 2012

Laboratory Investigation

105

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“…Conceptually, targeting the myoblast fusion pathway may represent a new avenue for PAX-FOXO1 RMS differentiation therapy, although whether an equivalent TANC1 axis participates in PAX-FOXO1-negative (i.e., embryonal) RMS remains a provocative question. Of note, Yang and colleagues have demonstrated that forced inhibition of MyoD in embryonal RMS cells prompts terminal differentiation (22). Therefore, RMS in general appears to be a clinically ripe candidate for differentiation therapy.…”

Section: Figurementioning

confidence: 99%

Drosophila and mammalian models uncover a role for the myoblast fusion gene TANC1 in rhabdomyosarcoma

Avirneni-Vadlamudi¹,

Galindo²,

Endicott³

et al. 2012

J. Clin. Invest.

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Rhabdomyosarcoma (RMS) is a malignancy of muscle myoblasts, which fail to exit the cell cycle, resist terminal differentiation, and are blocked from fusing into syncytial skeletal muscle. In some patients, RMS is caused by a translocation that generates the fusion oncoprotein PAX-FOXO1, but the underlying RMS pathogenetic mechanisms that impede differentiation and promote neoplastic transformation remain unclear. Using a Drosophila model of PAX-FOXO1-mediated transformation, we show here that mutation in the myoblast fusion gene rolling pebbles (rols) dominantly suppresses PAX-FOXO1 lethality. Further analysis indicated that PAX-FOXO1 expression caused upregulation of rols, which suggests that Rols acts downstream of PAX-FOXO1. In mammalian myoblasts, gene silencing of Tanc1, an ortholog of rols, revealed that it is essential for myoblast fusion, but is dispensable for terminal differentiation. Misexpression of PAX-FOXO1 in myoblasts upregulated Tanc1 and blocked differentiation, whereas subsequent reduction of Tanc1 expression to native levels by RNAi restored both fusion and differentiation. Furthermore, decreasing human TANC1 gene expression caused RMS cancer cells to lose their neoplastic state, undergo fusion, and form differentiated syncytial muscle. Taken together, these findings identify misregulated myoblast fusion caused by ectopic TANC1 expression as a RMS neoplasia mechanism and suggest fusion molecules as candidates for targeted RMS therapy.

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“…We first used human embryonal rhabdomyosarcoma cells (RD cells) 27,28 as a model system to study whether p53 has a direct role in regulating myogenic differentiation. RD cells carry a homozygous p53 Arg248Trp mutation, 29,30 which renders the protein defective in DNA binding.…”

Section: Resultsmentioning

confidence: 99%

“…The mouse MyoD gene was PCR amplified from pclBabe-MyoD. 27 Myogenin gene was PCR amplified from human cDNA prepared from RD cells. Lentiviral expression constructs, DHB-mVenus and H2B-mTurquoise, were kindly provided by Dr. Tobias Meyer.…”

Section: Methodsmentioning

confidence: 99%

p53 suppresses muscle differentiation at the myogenin step in response to genotoxic stress

et al. 2014

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Acute muscle injury and physiological stress from chronic muscle diseases and aging lead to impairment of skeletal muscle function. This raises the question of whether p53, a cellular stress sensor, regulates muscle tissue repair under stress conditions. By investigating muscle differentiation in the presence of genotoxic stress, we discovered that p53 binds directly to the myogenin promoter and represses transcription of myogenin, a member of the MyoD family of transcription factors that plays a critical role in driving terminal muscle differentiation. This reduction of myogenin protein is observed in G1-arrested cells and leads to decreased expression of late but not early differentiation markers. In response to acute genotoxic stress, p53-mediated repression of myogenin reduces post-mitotic nuclear abnormalities in terminally differentiated cells. This study reveals a mechanistic link previously unknown between p53 and muscle differentiation, and suggests new avenues for managing p53-mediated stress responses in chronic muscle diseases or during muscle aging. Cell Death and Differentiation (2015) 22, 560-573; doi:10.1038/cdd.2014; published online 12 December 2014The tumor suppressor p53 promotes cell cycle arrest or apoptosis in response to diverse stress signals such as DNA damage, thus preventing propagation of genetically compromised cells. [1][2][3] Among the diverse functions attributed to p53, a growing body of evidence supports its role in regulation of differentiation and maintenance of cellular function and integrity. 1,[4][5][6][7] For example, p53 represses Nanog to maintain genetic stability of the stem cell pool by promoting differentiation of mouse embryonic stem cells (mESCs) after DNA damage. 6 Skeletal muscle differentiation, a key step during muscle tissue formation, is orchestrated by the MyoD family of myogenic regulatory factors (MRFs). MyoD determines the myogenic lineage, whereas myogenin, a member of the MRF family, functions downstream of MyoD and plays a critical role in driving terminal differentiation as myogenin-null mice show a lethal deficiency of differentiated skeletal muscle. [8][9][10][11][12][13] The dynamic differentiation program of skeletal muscle is characterized by the orderly expression of genes and structural changes that can be recapitulated in vitro, as myogenic cells undergo cell cycle withdrawal and express early and then late differentiation genes with the formation of mononucleated myocytes to elongated multinucleated myotubes. 8,9,14 Under normal unstressed conditions, p53 has been shown to promote muscle differentiation in vitro. [15][16][17][18][19][20] In contrast, under stress-associated conditions such as inflammation, chronic exposure to double-strand DNA breaks, and aging, enhanced p53 activity has been shown to correlate with skeletal muscle atrophy in vivo. [21][22][23][24][25] In addition, it has been shown that p53 and its downstream effectors are required for an inflammatory cytokine-mediated inhibition of myogenic differentiation in vitro. 22 Int...

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MyoD and E-protein heterodimers switch rhabdomyosarcoma cells from an arrested myoblast phase to a differentiated state

Cited by 89 publications

References 58 publications

Downregulation of microRNAs miR-1, -206 and -29 stabilizes PAX3 and CCND2 expression in rhabdomyosarcoma

Downregulation of microRNAs miR-1, -206 and -29 stabilizes PAX3 and CCND2 expression in rhabdomyosarcoma

Drosophila and mammalian models uncover a role for the myoblast fusion gene TANC1 in rhabdomyosarcoma

p53 suppresses muscle differentiation at the myogenin step in response to genotoxic stress

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