Distinct Roles of the ISWI ATPases SMARCA1 and SMARCA5 in Fusion-Positive Rhabdomyosarcoma: A Division-of-Labor Hypothesis Linking Fusion Circuitry, Cell-State Plasticity, and Therapeutic Vulnerability

Fusion-positive rhabdomyosarcoma (FP-RMS) is driven by fusion transcription factors but retains an incomplete myogenic program. The roles of the two mammalian ISWI ATPases, SMARCA1/SNF2L and SMARCA5/SNF2H, remain poorly defined. This article proposes that they support FP-RMS through distinct but connected mechanisms. In RH30 cells, SMARCA1 loss markedly suppressed PAX3::FOXO1 and fusion-associated outputs, disrupted EGR1/Wnt- and TGF-β-linked chromatin programs, impaired differentiation and three-dimensional organization, and increased MEK-inhibitor sensitivity. SMARCA1 loss was also accompanied by marked depletion of MYCN and reductions in FUS and EP300, factors with established or emerging links to intrinsically disordered regions, transcriptional hubs, or biomolecular condensates. Together with recent evidence for PAX3::FOXO1/N-Myc transcriptional condensates, these observations motivate the hypothesis that SMARCA1-dependent chromatin competence may indirectly support the molecular environment required for fusion-associated hub organization, without implying that SMARCA1 itself nucleates condensates. By contrast, SMARCA5 was more closely associated with PI3K/mTOR signaling, cell-cycle progression, proliferation, and survival. In RH28 cells, strong SMARCA5 depletion with retained SMARCA1 reduced viability while preserving Wnt-responsive differentiation competence. Independent proximity labeling also detected SMARCA5 near selected FP-RMS fusion proteins, raising a context-specific possibility that SMARCA5 participates in selected fusion-associated molecular environments without establishing a condensate-scaffolding role. Single-cell analysis of 70,600 malignant cells from 16 RMS tumors further showed fusion-context-dependent associations of both ATPases with a high-risk transcriptional program. Together, these observations support a division-of-labor hypothesis in which SMARCA1 maintains chromatin competence for fusion-dependent and plastic cell states and may indirectly regulate fusion-associated transcriptional hubs, whereas SMARCA5 primarily supports proliferative fitness and may contribute to selected fusion-associated molecular environments. Matched perturbation, rescue, chromatin, protein, hub-imaging, single-cell, and three-dimensional studies are required to test this model.