Modeling early neuromuscular co-development: A 3D Neuromuscular organoids platform to unveil lncRNA dynamics in lineage bifurcation

Many neuromuscular disorders originate from defects during early embryogenesis, highlighting the critical need to understand the mechanisms governing neuro-muscular co-development. Because traditional 2D cultures fail to capture the spatial and cellular complexity of this cross-talk, advanced 3D human Neuro-Muscular Organoids (hNMOs) have emerged as an invaluable platform for modeling physiological co-development [1]. By combining this model system with single-cell RNA sequencing (scRNA-seq), we demonstrated that these organoids successfully give rise to all key lineage-specific subtypes - including muscle, neurons, glia, sclerotome, and epithelium - which collectively contribute to the proper development and formation of the muscle-nerve interaction. Within this framework, we identified highly promising, long noncoding RNA (lncRNA) candidates whose expression is tightly restricted to either the neural or mesodermal developmental trajectories. To dissect their functional relevance, we are generating specific lncRNA-knockout hiPSC lines, which will be used to establish KO-NMO models. Using these targeted organoid models, we will investigate how ablating cell-type specific lncRNAs alters early lineage commitment. Ultimately, we plan to elucidate the noncoding regulatory landscape underlying neuromuscular tissue assembly, with a particular focus on lncRNAs of interest in the lab, either muscle-specific [2], [3] or neural-enriched [4].