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A rapid one-generation genetic screen in a Drosophila model to capture rhabdomyosarcoma effectors and therapeutic targets.

Galindo KA, Endicott TR, Avirneni-Vadlamudi U, Galindo RL - G3 (Bethesda) (2014)

Bottom Line: Here, we report a new approach to dissect RMS, exploiting a highly efficient Drosophila PAX7-FOXO1 model uniquely configured to uncover PAX-FOXO1 RMS genetic effectors in only one generation.Additionally, we reveal that mutation of mastermind, a gene encoding a MEF2 transcriptional coactivator, similarly suppresses PAX7-FOXO1, further pointing toward MEF2 transcriptional activity as a PAX-FOXO1 underpinning.These studies show the utility of the PAX-FOXO1 Drosophila system as a robust one-generation (F1) RMS gene discovery platform and demonstrate how Drosophila transgenic conditional expression models can be configured for the rapid dissection of human disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390-9072.

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Mutation of mastermind, which encodes a MEF2 transcriptional cofactor, is a dominant PAX7-FOXO1 suppressor. (A) Overlapping chromosomal deletions identify a small genomic region, 50D1-50D5, as a PAX7-FOXO1−suppressing hotspot. (B) mastermind loss-of-function mutation dominantly suppresses PAX7-FOXO1 lethality. Df(2R)BSC18 was isolated in our original screen as a PAX7-FOXO1 suppressor (Table 1), which deletes mastermind (mam). The overlapping deletion, Df(2R)50C-36 also suppresses PAX7-FOXO1 (Table 2). Two well-characterized, strong loss-of-function mam alleles, mamBG02477 (n = 89 F1 adults scored) (P = 0.0044) and mam2 (n = 112 F1 adults scored) (P = 0.0043) suppress PAX7-FOXO1. Of note−although the mam2 allele showed a fold change of slightly less than 1.9, the increase in PAX7-FOXO1 adults (1.7-fold) was highly significant, and in this test we scored as a suppressor. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control.
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fig5: Mutation of mastermind, which encodes a MEF2 transcriptional cofactor, is a dominant PAX7-FOXO1 suppressor. (A) Overlapping chromosomal deletions identify a small genomic region, 50D1-50D5, as a PAX7-FOXO1−suppressing hotspot. (B) mastermind loss-of-function mutation dominantly suppresses PAX7-FOXO1 lethality. Df(2R)BSC18 was isolated in our original screen as a PAX7-FOXO1 suppressor (Table 1), which deletes mastermind (mam). The overlapping deletion, Df(2R)50C-36 also suppresses PAX7-FOXO1 (Table 2). Two well-characterized, strong loss-of-function mam alleles, mamBG02477 (n = 89 F1 adults scored) (P = 0.0044) and mam2 (n = 112 F1 adults scored) (P = 0.0043) suppress PAX7-FOXO1. Of note−although the mam2 allele showed a fold change of slightly less than 1.9, the increase in PAX7-FOXO1 adults (1.7-fold) was highly significant, and in this test we scored as a suppressor. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control.

Mentions: We next interrogated the 50D1-50D5 hotspot suppressor, which contains only five genes (Table 2 and Figure 5A), one of which is mam. In vivo studies in mammalian models have shown that the mam ortholog Mastermind-Like 1 (Maml1) encodes a transcriptional cofactor that physically interacts with Mef2 to augment Mef2-dependent promyogenic signaling (Shen et al. 2006; Potthoff and Olson 2007). Similar to D-Mef2, mam loss-of-function mutation dominantly suppressed PAX7-FOXO1−induced lethality (Figure 5B). Of note, mam expression levels were not detectably altered in our PAX7-FOXO1 microarray studies, compatible with mam’s role as a cofactor vs. myogenesis gene target. Taken together, these Drosophila studies highlight a putative PAX-FOXO1→MEF2→RMS pathogenic axis, while also demonstrating that the one-generation (F1) genetic screen quickly uncovers dominant PAX-FOXO1 modifiers/effectors in an unbiased fashion.


A rapid one-generation genetic screen in a Drosophila model to capture rhabdomyosarcoma effectors and therapeutic targets.

Galindo KA, Endicott TR, Avirneni-Vadlamudi U, Galindo RL - G3 (Bethesda) (2014)

Mutation of mastermind, which encodes a MEF2 transcriptional cofactor, is a dominant PAX7-FOXO1 suppressor. (A) Overlapping chromosomal deletions identify a small genomic region, 50D1-50D5, as a PAX7-FOXO1−suppressing hotspot. (B) mastermind loss-of-function mutation dominantly suppresses PAX7-FOXO1 lethality. Df(2R)BSC18 was isolated in our original screen as a PAX7-FOXO1 suppressor (Table 1), which deletes mastermind (mam). The overlapping deletion, Df(2R)50C-36 also suppresses PAX7-FOXO1 (Table 2). Two well-characterized, strong loss-of-function mam alleles, mamBG02477 (n = 89 F1 adults scored) (P = 0.0044) and mam2 (n = 112 F1 adults scored) (P = 0.0043) suppress PAX7-FOXO1. Of note−although the mam2 allele showed a fold change of slightly less than 1.9, the increase in PAX7-FOXO1 adults (1.7-fold) was highly significant, and in this test we scored as a suppressor. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4321029&req=5

fig5: Mutation of mastermind, which encodes a MEF2 transcriptional cofactor, is a dominant PAX7-FOXO1 suppressor. (A) Overlapping chromosomal deletions identify a small genomic region, 50D1-50D5, as a PAX7-FOXO1−suppressing hotspot. (B) mastermind loss-of-function mutation dominantly suppresses PAX7-FOXO1 lethality. Df(2R)BSC18 was isolated in our original screen as a PAX7-FOXO1 suppressor (Table 1), which deletes mastermind (mam). The overlapping deletion, Df(2R)50C-36 also suppresses PAX7-FOXO1 (Table 2). Two well-characterized, strong loss-of-function mam alleles, mamBG02477 (n = 89 F1 adults scored) (P = 0.0044) and mam2 (n = 112 F1 adults scored) (P = 0.0043) suppress PAX7-FOXO1. Of note−although the mam2 allele showed a fold change of slightly less than 1.9, the increase in PAX7-FOXO1 adults (1.7-fold) was highly significant, and in this test we scored as a suppressor. *P < 0.05, **P < 0.01, ***P < 0.001 vs. control.
Mentions: We next interrogated the 50D1-50D5 hotspot suppressor, which contains only five genes (Table 2 and Figure 5A), one of which is mam. In vivo studies in mammalian models have shown that the mam ortholog Mastermind-Like 1 (Maml1) encodes a transcriptional cofactor that physically interacts with Mef2 to augment Mef2-dependent promyogenic signaling (Shen et al. 2006; Potthoff and Olson 2007). Similar to D-Mef2, mam loss-of-function mutation dominantly suppressed PAX7-FOXO1−induced lethality (Figure 5B). Of note, mam expression levels were not detectably altered in our PAX7-FOXO1 microarray studies, compatible with mam’s role as a cofactor vs. myogenesis gene target. Taken together, these Drosophila studies highlight a putative PAX-FOXO1→MEF2→RMS pathogenic axis, while also demonstrating that the one-generation (F1) genetic screen quickly uncovers dominant PAX-FOXO1 modifiers/effectors in an unbiased fashion.

Bottom Line: Here, we report a new approach to dissect RMS, exploiting a highly efficient Drosophila PAX7-FOXO1 model uniquely configured to uncover PAX-FOXO1 RMS genetic effectors in only one generation.Additionally, we reveal that mutation of mastermind, a gene encoding a MEF2 transcriptional coactivator, similarly suppresses PAX7-FOXO1, further pointing toward MEF2 transcriptional activity as a PAX-FOXO1 underpinning.These studies show the utility of the PAX-FOXO1 Drosophila system as a robust one-generation (F1) RMS gene discovery platform and demonstrate how Drosophila transgenic conditional expression models can be configured for the rapid dissection of human disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390-9072.

Show MeSH
Related in: MedlinePlus