Single-nucleus RNA-seq identifies divergent populations of FSHD2 myotube nuclei

by Shan Jiang, Katherine Williams, Xiangduo Kong, Weihua Zeng, Nam Viet Nguyen, Xinyi Ma, Rabi Tawil, Kyoko Yokomori, Ali Mortazavi

FSHD is characterized by the misexpression ofDUX4 in skeletal muscle. AlthoughDUX4 upregulation is thought to be the pathogenic cause of FSHD,DUX4 is lowly expressed in patient samples, and analysis of the consequences ofDUX4 expression has largely relied on artificial overexpression. To better understand the native expression profile ofDUX4 and its targets, we performed bulk RNA-seq on a 6-day differentiation time-course in primary FSHD2 patient myoblasts. We identify a set of 54 genes upregulated in FSHD2 cells, termed FSHD-induced genes. Using single-cell and single-nucleus RNA-seq on myoblasts and differentiated myotubes, respectively, we captured, for the first time,DUX4 expressed at the single-nucleus level in a native state. We identified two populations of FSHD myotube nuclei based on low or high enrichment ofDUX4 and FSHD-induced genes ( “FSHD-Lo” and “FSHD Hi”, respectively). FSHD-Hi myotube nuclei coexpress multiple DUX4 target genes includingDUXA,LEUTX andZSCAN4, and also upregulate cell cycle-related genes with significant enrichment of E2F target genes and p53 signaling activation. We found more FSHD-Hi nuclei thanDUX4-positive nuclei, and confirmed within situ RNA/protein detection thatDUX4 transcribed in only one or two nuclei is sufficient for DUX4 protein to activate target genes across multiple nuclei within the same m...

Source: PLoS Genetics - May 3, 2020 Category: Genetics & Stem Cells Authors: Shan Jiang Source Type: research