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  3. Acetylation profiling by Iseq-Kac reveals insights into HSC aging and lineage decision

Acetylation profiling by Iseq-Kac reveals insights into HSC aging and lineage decision

  • Nat Chem Biol. 2025 May 26. doi: 10.1038/s41589-025-01916-1.
Yanqiu Gong # 1,  Huiwen Zhan # 1,  Ni Wei # 1,  Min Liu # 1,  Yu Liu # 1,  Pengbo Guan 1,  Yusi Xie 1,  Yujun Deng 1,  Qianlun Pu 1,  Xiaoxian Lou 1,  Xiaodong Wang 1,  Rou Zhang 1,  Ping Wang 2,  Xiuxiu Jin 1,  Xiuxuan Wang 1,  Zhiqiang Xu 1,  Li Gao 1,  Xinyuan Wang 1,  Siyu He 1,  Ying Lu 1,  Meng Hu 1,  Wanmeng Li 1,  Kun Zheng 1,  Yong Peng 1,  Peng Lei 1,  Heng Xu 1,  Yujun Shi 3,  Jun Qin 4,  Hongbo Hu 5 6 7,  Huiyuan Zhang 8,  Lunzhi Dai 9 10
Affiliations

Affiliations

  • 1 Center for Hematology and Immunology and National Clinical Research Center for Geriatrics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China.
  • 2 School of Medical Imaging, Tianjin Medical University, Tianjin, China.
  • 3 Laboratory of Pathology, Key Laboratory of Transplant Engineering and Immunology, NHC, West China Hospital, Sichuan University, Chengdu, China.
  • 4 State Key Laboratory of Proteomics, Institute of Lifeomics, Beijing, China.
  • 5 Center for Hematology and Immunology and National Clinical Research Center for Geriatrics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China. hongbohu@scu.edu.cn.
  • 6 Chongqing International Institute for Immunology, Chongqing, China. hongbohu@scu.edu.cn.
  • 7 Tianfu Jincheng Laboratory, Chengdu, China. hongbohu@scu.edu.cn.
  • 8 Center for Hematology and Immunology and National Clinical Research Center for Geriatrics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China. hyzhang@scu.edu.cn.
  • 9 Center for Hematology and Immunology and National Clinical Research Center for Geriatrics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China. lunzhi.dai@scu.edu.cn.
  • 10 Tianfu Jincheng Laboratory, Chengdu, China. lunzhi.dai@scu.edu.cn.
  • # Contributed equally.
PMID: 40419771 DOI: 10.1038/s41589-025-01916-1
Abstract

Profiling post-translational modifications face challenges with low-input samples. We developed Iseq-Kac (internal standard-assisted enrichment-free approach for high-throughput quantitative analysis of lysine acetylation) to profile the acetylome in as few as 103-104 cells. By using a hyperacetylated internal standard, Iseq-Kac can be used in mass spectrometry (MS) to enhance MS1 signals and facilitate MS2 fragmentation of acetylated peptides. Using Iseq-Kac, we quantified 675-1,471 acetylated peptides per analysis from 104 hematopoietic stem cells (HSCs) or multipotent progenitors. Validation by targeted MS, site-specific antibodies and functional assays linked aging-related proteome and acetylome changes to HSC lineage decision. A pronounced decrease in acetylation at H4 lysine 77 (H4K77ac) was observed in aged HSCs, linked to histone deacetylase 3 (HDAC3) activity. HDAC3 inhibition or knockdown in HSCs significantly promoted lymphocyte differentiation. Mimicking H4K77ac through H4K77Q expression enhanced B cell differentiation while repressing myeloid differentiation. Overall, Iseq-Kac enables robust low-input acetylome profiling and reveals epigenetic mechanisms underlying lineage skewing in aged HSCs.

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