Single-nucleus RNA-seq identifies divergent populations of FSHD2 myotube nuclei.
Case-Control Studies
Cell Differentiation
Cell Nucleus
/ chemistry
Cells, Cultured
Gene Expression Regulation
HEK293 Cells
Humans
Microfilament Proteins
/ genetics
Muscle Fibers, Skeletal
/ metabolism
Muscular Dystrophy, Facioscapulohumeral
/ genetics
Myoblasts
/ metabolism
Nuclear Proteins
/ genetics
RNA-Binding Proteins
/ genetics
RNA-Seq
/ methods
Single-Cell Analysis
/ methods
Exome Sequencing
Journal
PLoS genetics
ISSN: 1553-7404
Titre abrégé: PLoS Genet
Pays: United States
ID NLM: 101239074
Informations de publication
Date de publication:
05 2020
05 2020
Historique:
received:
27
02
2019
accepted:
03
04
2020
revised:
14
05
2020
pubmed:
5
5
2020
medline:
31
7
2020
entrez:
5
5
2020
Statut:
epublish
Résumé
FSHD is characterized by the misexpression of DUX4 in skeletal muscle. Although DUX4 upregulation is thought to be the pathogenic cause of FSHD, DUX4 is lowly expressed in patient samples, and analysis of the consequences of DUX4 expression has largely relied on artificial overexpression. To better understand the native expression profile of DUX4 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 of DUX4 and FSHD-induced genes ("FSHD-Lo" and "FSHD Hi", respectively). FSHD-Hi myotube nuclei coexpress multiple DUX4 target genes including DUXA, LEUTX and ZSCAN4, and also upregulate cell cycle-related genes with significant enrichment of E2F target genes and p53 signaling activation. We found more FSHD-Hi nuclei than DUX4-positive nuclei, and confirmed with in situ RNA/protein detection that DUX4 transcribed in only one or two nuclei is sufficient for DUX4 protein to activate target genes across multiple nuclei within the same myotube. DUXA (the DUX4 paralog) is more widely expressed than DUX4, and depletion of DUXA suppressed the expression of LEUTX and ZSCAN4 in late, but not early, differentiation. The results suggest that the DUXA can take over the role of DUX4 to maintain target gene expression. These results provide a possible explanation as to why it is easier to detect DUX4 target genes than DUX4 itself in patient cells and raise the possibility of a self-sustaining network of gene dysregulation triggered by the limited DUX4 expression.
Identifiants
pubmed: 32365093
doi: 10.1371/journal.pgen.1008754
pii: PGENETICS-D-19-00284
pmc: PMC7224571
doi:
Substances chimiques
FRG1 protein, human
0
FRG2 protein, human
0
Microfilament Proteins
0
Nuclear Proteins
0
RNA-Binding Proteins
0
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
e1008754Subventions
Organisme : NIAMS NIH HHS
ID : R21 AR071104
Pays : United States
Déclaration de conflit d'intérêts
The authors have declared that no competing interests exist.
Références
Hum Mol Genet. 2016 Oct 15;25(20):4419-4431
pubmed: 28171552
Muscle Nerve. 2006 Jul;34(1):1-15
pubmed: 16508966
Nat Rev Cancer. 2013 Aug;13(8):585-95
pubmed: 23842645
Nucleic Acids Res. 2009 May;37(9):e68
pubmed: 19357094
Hum Mol Genet. 2019 Apr 1;28(7):1064-1075
pubmed: 30445587
Science. 2010 Sep 24;329(5999):1650-3
pubmed: 20724583
Ann Neurol. 2011 Mar;69(3):540-52
pubmed: 21446026
Nucleic Acids Res. 2016 Dec 1;44(21):e158
pubmed: 27566152
Int J Mol Sci. 2017 Oct 19;18(10):
pubmed: 29048367
Mol Cell. 2010 May 28;38(4):576-89
pubmed: 20513432
Bioinformatics. 2013 Jan 1;29(1):15-21
pubmed: 23104886
Hum Mutat. 2014 Aug;35(8):998-1010
pubmed: 24838473
PLoS One. 2011;6(10):e26820
pubmed: 22053214
Hum Mol Genet. 2015 Oct 15;24(20):5901-14
pubmed: 26246499
J Cell Mol Med. 2013 Jan;17(1):76-89
pubmed: 23206257
Trends Genet. 2017 Apr;33(4):233-243
pubmed: 28222895
Nat Protoc. 2014 Jan;9(1):171-81
pubmed: 24385147
PLoS Comput Biol. 2010 Jul 15;6(7):e1000852
pubmed: 20657661
Dev Cell. 2012 Jan 17;22(1):38-51
pubmed: 22209328
Am J Hum Genet. 2013 Oct 3;93(4):744-51
pubmed: 24075187
Genome Biol. 2019 Dec 23;20(1):296
pubmed: 31870423
Hum Mol Genet. 2019 Apr 15;28(8):1244-1259
pubmed: 30462217
Nucleic Acids Res. 2019 Jan 8;47(D1):D33-D38
pubmed: 30204897
Nat Genet. 2017 Jun;49(6):941-945
pubmed: 28459456
Genome Res. 2013 Aug;23(8):1195-209
pubmed: 23595228
Bioinformatics. 2016 Sep 15;32(18):2847-9
pubmed: 27207943
Genome Res. 2003 Nov;13(11):2498-504
pubmed: 14597658
J Cell Sci. 2016 Oct 15;129(20):3816-3831
pubmed: 27744317
Nat Biotechnol. 2018 Jun;36(5):411-420
pubmed: 29608179
Nat Genet. 2003 Dec;35(4):315-7
pubmed: 14634647
BMC Bioinformatics. 2011 Aug 04;12:323
pubmed: 21816040
Nucleic Acids Res. 2018 Jan 4;46(D1):D252-D259
pubmed: 29140464
Bioinformatics. 2010 Jan 1;26(1):139-40
pubmed: 19910308
BMC Evol Biol. 2010 Nov 26;10:364
pubmed: 21110847
Nat Genet. 2017 Jun;49(6):935-940
pubmed: 28459454
Nat Genet. 2017 Jun;49(6):925-934
pubmed: 28459457
PLoS Genet. 2013 Nov;9(11):e1003947
pubmed: 24278031
BMC Genomics. 2018 Dec 24;19(1):960
pubmed: 30583719
Elife. 2015 Jan 07;4:
pubmed: 25564732
EMBO J. 2008 Oct 22;27(20):2766-79
pubmed: 18833193
PLoS Genet. 2009 Jul;5(7):e1000559
pubmed: 19593370
PLoS Genet. 2010 Oct 28;6(10):e1001181
pubmed: 21060811
Hum Mol Genet. 2014 Oct 15;23(20):5342-52
pubmed: 24861551
Nat Commun. 2019 Apr 3;10(1):1523
pubmed: 30944313
Antioxid Redox Signal. 2015 Jun 1;22(16):1463-82
pubmed: 25336259
Nat Genet. 2012 Dec;44(12):1370-4
pubmed: 23143600
Cell Rep. 2017 Feb 14;18(7):1713-1726
pubmed: 28199843
Elife. 2018 Mar 13;7:
pubmed: 29533181
Cell Rep. 2019 Nov 12;29(7):1812-1820.e5
pubmed: 31722199
Eur J Hum Genet. 2015 Jun;23(6):808-16
pubmed: 25370034