Single-nucleus RNA-seq identifies transcriptional heterogeneity in multinucleated skeletal myofibers

While the majority of cells contain a single nucleus, cell types such as trophoblasts, osteoclasts, and skeletal myofibers require multinucleation. One advantage of multinucleation can be the assignment of distinct functions to different nuclei, but comprehensive interrogation of transcriptional heterogeneity within multinucleated tissues has been challenging due to the presence of a shared cytoplasm. Here, we utilized single-nucleus RNA-sequencing (snRNA-seq) to determine the extent of transcriptional diversity within multinucleated skeletal myofibers. Nuclei from mouse skeletal muscle were profiled across the lifespan, which revealed the emergence of distinct myonuclear populations in postnatal development and their reactivation in aging muscle. Our datasets also provided a platform for discovery of novel genes associated with rare specialized regions of the muscle cell, including markers of the myotendinous junction and functionally validated factors expressed at the neuromuscular junction. These findings reveal that myonuclei within syncytial muscle fibers possess distinct transcriptional profiles that regulate muscle biology. Single-nucleus RNA-seq of tibialis anteior muscle of BL6 wild-type mice across the lifespan: 10 days, 21 days, 5 months (TA and soleus), 24 months, 30 months尽管绝大多数细胞仅含单个细胞核，但滋养层细胞（trophoblasts）、破骨细胞（osteoclasts）与骨骼肌肌纤维（skeletal myofibers）等细胞类型则需形成多核结构。多核化的优势之一，在于可使不同细胞核承担差异化功能，但由于多核组织共享细胞质，对其转录异质性开展全面解析始终颇具挑战。本研究借助单细胞核RNA测序（single-nucleus RNA-sequencing, snRNA-seq）技术，解析了多核骨骼肌肌纤维内的转录多样性程度。我们对小鼠骨骼肌的细胞核进行了全生命周期的转录组表征，结果揭示出生后发育阶段中独特肌细胞核群体的形成，以及衰老肌肉中这些群体的重新激活。本研究数据集还为发现肌细胞罕见特化区域相关的新型基因提供了支撑平台，其中涵盖肌腱连接（myotendinous junction）标志物，以及在神经肌肉接头（neuromuscular junction）处表达并经功能验证的调控因子。上述研究结果证实，合胞体肌纤维内的肌细胞核拥有调控肌肉生物学功能的独特转录谱。本数据集为BL6野生型小鼠全生命周期胫骨前肌（tibialis anterior）的单细胞核RNA测序数据，涵盖时间点包括：出生后10天、21天、5月龄（含胫骨前肌与比目鱼肌）、24月龄及30月龄。