SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes

Okuno, Shota; Higo, Shuichiro; Kondo, Takumi; Shiba, Mikio; Kameda, Satoshi; Inoue, Hiroyuki; Tabata, Tomoka; Ogawa, Shou; Morishita, Yu; Sun, Congcong; Ishino, Saki; Honda, Tomoyuki; Miyagawa, Shigeru; Sakata, Yasushi

doi:https://doi.org/10.1038/s41598-023-48084-7

SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes

Okuno, Shota; Higo, Shuichiro; Kondo, Takumi et al.

Scientific Reports, 2023, 13(1), 21397

Number of Access:208（2025-06-03 00:59 Counts）

Identifier to cite or link to this item: https://hdl.handle.net/11094/93523

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Title	SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes
Creator	CreatorName Okuno, Shota
	NRID 1000000604034 1000000604034 researchmap permalink s.higo s.higo Scopus Author ID 23018633200 23018633200 CreatorName Higo, Shuichiro
	CreatorName Kondo, Takumi
	CreatorName Shiba, Mikio
	CreatorName Kameda, Satoshi
	CreatorName Inoue, Hiroyuki
	NRID 1000030973094 1000030973094 researchmap permalink tomoka0122 tomoka0122 CreatorName Tabata, Tomoka
	CreatorName Ogawa, Shou
	CreatorName Morishita, Yu
	CreatorName Sun, Congcong
	CreatorName Ishino, Saki
	NRID 1000080402676 1000080402676 CreatorName Honda, Tomoyuki
	NRID 1000070544237 1000070544237 ORCID 0000-0003-0015-6569 0000-0003-0015-6569 researchmap permalink s-miyagawa3154 s-miyagawa3154 Scopus Author ID 55560624700 55560624700 CreatorName Miyagawa, Shigeru
	NRID 1000000397671 1000000397671 ORCID 0000-0002-5618-4721 0000-0002-5618-4721 researchmap permalink C4589 C4589 Scopus Author ID 35395964700 35395964700 CreatorName Sakata, Yasushi
Description	Okuno S., Higo S., Kondo T., et al. SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes. Scientific Reports 13, 21397 (2023); https://doi.org/10.1038/s41598-023-48084-7.
Abstract	Although an increased risk of myocarditis has been observed after vaccination with mRNA encoding severe acute respiratory syndrome coronavirus 2 spike protein, its underlying mechanism has not been elucidated. This study investigated the direct effects of spike receptor-binding domain (S-RBD) on human cardiomyocytes differentiated from induced pluripotent stem cells (iPSC-CMs). Immunostaining experiments using ACE2 wild-type (WT) and knockout (KO) iPSC-CMs treated with purified S-RBD demonstrated that S-RBD was bound to ACE2 and internalized into the subcellular space in the iPSC-CMs, depending on ACE2. Immunostaining combined with live cell imaging using a recombinant S-RBD fused to the superfolder GFP (S-RBD-sfGFP) demonstrated that S-RBD was bound to the cell membrane, co-localized with RAB5A, and then delivered from the endosomes to the lysosomes in iPSC-CMs. Quantitative PCR array analysis followed by single cell RNA sequence analysis clarified that S-RBD-sfGFP treatment significantly upregulated the NF-kβ pathway-related gene (CXCL1) in the differentiated non-cardiomyocytes, while upregulated interferon (IFN)-responsive genes (IFI6, ISG15, and IFITM3) in the matured cardiomyocytes. S-RBD-sfGFP treatment promoted protein ISGylation, an ISG15-mediated post-translational modification in ACE2-WT-iPSC-CMs, which was suppressed in ACE2-KO-iPSC-CMs. Our experimental study demonstrates that S-RBD is internalized through the endolysosomal pathway, which upregulates IFN-responsive genes and promotes ISGylation in the iPSC-CMs.
Publisher	Nature Research
Source Title	Scientific Reports
Volume (Issue)	13(1)
Page	21397
Date of Issued	2023-12-04
Language	English
Handle URL	https://hdl.handle.net/11094/93523
eISSN	20452322
Relation.isIdenticalTo	DOI (Publisher Version) https://doi.org/10.1038/s41598-023-48084-7
Access Rights	open access
CopyRight	This article is licensed under a Creative Commons Attribution 4.0 International License.
oaire:version	Version of Record
Category	学術雑誌論文 Journal Article

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Resource Type	journal article
Local Type	Journal Article
DCMI Type	Text
DCTERMS.bibliographicCitation	Scientific Reports. 2023, 13(1), p. 21397
DC.title	SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes
DC.creator	Okuno, Shota
	Higo, Shuichiro
	Kondo, Takumi
	Shiba, Mikio
	Kameda, Satoshi
	Inoue, Hiroyuki
	Tabata, Tomoka
	Ogawa, Shou
	Morishita, Yu
	Sun, Congcong
	Ishino, Saki
	Honda, Tomoyuki
	Miyagawa, Shigeru
	Sakata, Yasushi
DC.publisher	Nature Research
DC.language' scheme='DCTERMS.RFC1766	English
DC.type' scheme='DCTERMS.DCMIType	Text
DCTERMS.issued' scheme='DCTERMS.W3CDTF	2023-12-04
DC.identifier	https://doi.org/10.1038/s41598-023-48084-7
DC.description	Okuno S., Higo S., Kondo T., et al. SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes. Scientific Reports 13, 21397 (2023); https://doi.org/10.1038/s41598-023-48084-7.
DCTERMS.abstract	Although an increased risk of myocarditis has been observed after vaccination with mRNA encoding severe acute respiratory syndrome coronavirus 2 spike protein, its underlying mechanism has not been elucidated. This study investigated the direct effects of spike receptor-binding domain (S-RBD) on human cardiomyocytes differentiated from induced pluripotent stem cells (iPSC-CMs). Immunostaining experiments using ACE2 wild-type (WT) and knockout (KO) iPSC-CMs treated with purified S-RBD demonstrated that S-RBD was bound to ACE2 and internalized into the subcellular space in the iPSC-CMs, depending on ACE2. Immunostaining combined with live cell imaging using a recombinant S-RBD fused to the superfolder GFP (S-RBD-sfGFP) demonstrated that S-RBD was bound to the cell membrane, co-localized with RAB5A, and then delivered from the endosomes to the lysosomes in iPSC-CMs. Quantitative PCR array analysis followed by single cell RNA sequence analysis clarified that S-RBD-sfGFP treatment significantly upregulated the NF-kβ pathway-related gene (CXCL1) in the differentiated non-cardiomyocytes, while upregulated interferon (IFN)-responsive genes (IFI6, ISG15, and IFITM3) in the matured cardiomyocytes. S-RBD-sfGFP treatment promoted protein ISGylation, an ISG15-mediated post-translational modification in ACE2-WT-iPSC-CMs, which was suppressed in ACE2-KO-iPSC-CMs. Our experimental study demonstrates that S-RBD is internalized through the endolysosomal pathway, which upregulates IFN-responsive genes and promotes ISGylation in the iPSC-CMs.
DC.rights	This article is licensed under a Creative Commons Attribution 4.0 International License.
DC.format	93523
citation_title	SARS-CoV-2 spike receptor-binding domain is internalized and promotes protein ISGylation in human induced pluripotent stem cell-derived cardiomyocytes
citation_author	Okuno, Shota
	Higo, Shuichiro
	Kondo, Takumi
	Shiba, Mikio
	Kameda, Satoshi
	Inoue, Hiroyuki
	Tabata, Tomoka
	Ogawa, Shou
	Morishita, Yu
	Sun, Congcong
	Ishino, Saki
	Honda, Tomoyuki
	Miyagawa, Shigeru
	Sakata, Yasushi
citation_publisher	Nature Research
citation_language	English
citation_date	2023-12-04
citation_journal_title	Scientific Reports
citation_volume	13
citation_issue	1
citation_firstpage	21397
citation_doi	https://doi.org/10.1038/s41598-023-48084-7
citation_public_url	https://hdl.handle.net/11094/93523