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Status |
Public on May 19, 2024 |
Title |
Histone lactylation-mediated NSUN2 transcriptional activation dictates choroidal neovascularization through promoting AKAP2 mRNA expression [CUT&Tag] |
Organism |
Homo sapiens |
Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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Summary |
Background: As a widespread post-transcriptional RNA modification, N5-methylcytosine (m5C) is implicated in a variety of cellular responses and processes that regulate RNA metabolism. Despite this, a clear understanding of m5C modification’s role and mechanism in angiogenesis is still lacking. Methods: Single-cell RNA sequencing data was analyzed to determine expression of m5C methylase NSUN2. m5C levels were determined by mRNA isolation and anti-m5C dot blot in both hypoxia-induced endothelial cells (ECs) and laser-induced choroidal neovascularization (CNV). In addition, endothelial cell and endothelium‐specific NSUN2‐knockout mouse model were used to investigate the effect of NSUN2 silence on angiogenic phenotype. Genome-wide multiomics analyses were performed to identify the functional target of NSUN2, including proteomic analysis, transcriptome screening and m5C-methylated RNA immunoprecipitation sequencing (m5C-meRIP-seq). CUT&Tag sequencing was performed to test the histone lactylation signal in the promoter region of NSUN2. Finally, AAV-mediated short hairpin RNAi knockdown of NSUN2 gene expression (AAV-shNSUN2) was constructed to investigate the effect of inhibiting CNV. Results: First, we discovered that m5C methylase NSUN2 expression level and mRNA m5C level were significantly higher in CNV-ECs than in normal ECs. NSUN2 knockdown in ECs inhibited proliferative, migration, and tube formation activities of ECs. Moreover, compared with EC NSUN2flox/flox mice, EC-specific NSUN2-deficient (EC NSUN2-/-) mice displayed less retinal vascular leakage after laser induction. Through multiomics analyses, we subsequently identified A-kinase anchoring protein 2 (AKAP2), a scaffolding protein which isolate Protein kinase A (PKA) to specific subcellular locations through binding to its regulatory subunit, as a downstream candidate target of NSUN2 in ECs. Overexpression of exogenous AKAP2 markedly reversed the inhibitory phenotypes in NSUN2-deficient ECs. Interestingly, laser induced NSUN2 up-regulation was driven by lactate-mediated lactylation on histone H3K18. In CNV models, AAV-mediated repression of NSUN2 modulated highly retinal vascular leakage and choroidal thickness. Conclusion: Overall, our findings indicate that NSUN2 is a novel therapeutic target for choroidal neovascularization.
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Overall design |
CUT&Tag-seq data related to gene NSUN2 in human umbilical vein endothelial cell
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Contributor(s) |
Zuo S, Li L, Lu L, Fan X |
Citation missing |
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Submission date |
May 16, 2024 |
Last update date |
May 19, 2024 |
Contact name |
Sipeng Zuo |
E-mail(s) |
Zsp2019@sjtu.edu.cn
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Organization name |
Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine
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Street address |
No. 12, Lane 833, Zhizaoju Road, Huangpu District
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City |
Shanghai |
State/province |
Shanghai |
ZIP/Postal code |
200023 |
Country |
China |
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Platforms (1) |
GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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Samples (4)
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Relations |
BioProject |
PRJNA1112365 |
Supplementary file |
Size |
Download |
File type/resource |
GSE267661_All.xls.gz |
217 b |
(ftp)(http) |
XLS |
SRA Run Selector |
Raw data are available in SRA |
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