Visualization Imaging of Cell Membrane Receptor Proteins Based on Non-genetic Engineering Approaches

doi:10.6023/A24120363

Acta Chimica Sinica ›› 2025, Vol. 83 ›› Issue (3): 299-308.DOI: 10.6023/A24120363 Previous Articles Next Articles

Review

基于非基因工程的细胞膜受体蛋白可视化成像

周淼淼^a, 曹浩文^a, 杨心怡^a, 任心怡^a, 姚峰^a^,^b^,^*(), 何磊良^a^,^*()

a 郑州大学公共卫生学院郑州 450001
b 中国铁路南昌局集团有限公司南昌疾病预防控制所南昌 330003

投稿日期:2024-12-03 发布日期:2025-01-23

作者简介:

周淼淼, 2000年9月出生, 安徽阜阳人, 自2022年9月起加入郑州大学公共卫生学院何磊良课题组从事硕士研究工作. 主要研究方向涉及细胞膜蛋白靶向降解.

曹浩文, 2002年7月出生, 安徽阜阳人, 自2024年9月起加入郑州大学公共卫生学院何磊良课题组从事硕士研究工作, 主要研究方向涉及核酸探针设计与生物传感.

杨心怡, 2001年10月出生, 山东潍坊人, 自2024年9月起加入郑州大学公共卫生学院何磊良课题组从事硕士研究工作, 主要研究方向涉及核酸探针设计与生物传感.

任心怡, 2002年12月出生, 山西汾阳人, 自2024年9月起加入郑州大学公共卫生学院何磊良课题组从事硕士研究工作, 主要研究方向涉及核酸探针设计与生物传感.

姚峰, 1997年10月出生, 江西宜春人, 自2021年9月起加入郑州大学公共卫生学院何磊良课题组从事硕士研究工作, 主要研究方向涉及细胞膜蛋白寡聚化成像.

何磊良, 教授, 2015年于湖南大学获博士学位, 何磊良教授目前在郑州大学公共卫生学院进行研究工作, 主要研究兴趣包括肿瘤液体活检及单细胞分析与调控.

基金资助:
国家自然科学基金(82373630); 国家自然科学基金(82073606); 河南省本科高校青年骨干教师项目(2023GGJS009)

Visualization Imaging of Cell Membrane Receptor Proteins Based on Non-genetic Engineering Approaches

Miaomiao Zhou^a, Haowen Cao^a, Xinyi Yang^a, Xinyi Ren^a, Feng Yao^a^,^b(), Leiliang He^a()

a College of Public Health, Zhengzhou University, Zhengzhou 450001, China
b Nanchang Disease Prevention and Control Institute, China Railway Nanchang Bureau Group Co., Ltd., Nanchang 330003, China

Received:2024-12-03 Published:2025-01-23
Contact: ^*E-mail: yaofeng202166@163.com; hell2015@zzu.edu.cn
Supported by:
National Natural Science Foundations of China(82373630); National Natural Science Foundations of China(82073606); training grant of Henan Province for Young Backbone Teachers(2023GGJS009)

Cell membrane receptor proteins are important mediators for communication between cells and the external environment. Changes in their expression levels or behavior may indicate the initiation of certain physiological or pathological processes in the body, and many cell membrane receptor proteins have been used as a basis for identifying different cell populations or subtypes and as targets for targeted therapy. Traditional genetic engineering protein imaging strategies involve genetic manipulation, which may interfere with other biological processes within the cell and have unpredictable limitations. Therefore, non-genetic engineering protein imaging strategies have gained significant attention in tumor visualization imaging research, mainly due to their notable characteristics, including simplicity, programmable design, and ease of regulation, and have been widely used in the imaging studies of cell membrane receptor proteins. This review comprehensively summarizes the use of non-genetic engineering strategies for cell membrane receptor protein imaging. It includes the design of logical gate probes, fluorescence resonance energy transfer (FRET) probes, structurally constrained hybridization probes, and signal tags for cell type identification; secondly, it summarizes the application of this protein imaging mode in the interaction of cell membrane receptor proteins, where receptor dimerization in the interaction of membrane receptor proteins is the first step in receptor activation and cell communication, and the visualization of membrane receptor dimerization is achieved by designing DNA probe systems and fluorescent sensors. Additionally, this review covers the application of spatial distribution analysis imaging of cell membrane receptor proteins, achieving nanometer-level spatial distribution imaging of membrane receptors through single-molecule localization microscopy (SMLM) and point accumulation for imaging in nanoscale topography (PAINT); finally, it looks forward to the challenges faced in this field and future development directions. With advancements in DNA nanotechnology, imaging technology, and bioinformatics, future research on non-genetic engineering strategies could provide higher resolution and more in-depth imaging of cell membrane receptor proteins, thereby offering more innovative pathways for in-depth study of intercellular communication mechanisms.

Key words: cell membrane receptor proteins, non-genetic engineering, visualization imaging, cell type identification, receptor dimerization, spatial distribution analysis

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Miaomiao Zhou, Haowen Cao, Xinyi Yang, Xinyi Ren, Feng Yao, Leiliang He. Visualization Imaging of Cell Membrane Receptor Proteins Based on Non-genetic Engineering Approaches[J]. Acta Chimica Sinica, 2025, 83(3): 299-308.

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Fig. & Tab. 6

Figure 1. Logical gate-based recognition of membrane receptors for cell type identification (A) A mechanism of associative toehold activation based on multiple aptamers for logical operations to identify different cancer cell subtypes[32]; (B) A DNA logic gate nanorobot (DNA-TP) based on dual aptamers (sgc8c/sgc4f) as recognition molecules, achieving precise cancer cell identification through dual-specificity recognition and logical operations[33]; (C) A DNA nanorobot (pH-RE) constructed from programmable modules that simultaneously recognizes extracellular pH and cancer cell membrane receptors[35]

Figure 2. FRET for membrane receptor recognition in cell type identification (A) Hybridization chain reaction (HCR) amplification and FRET signal amplification for the visualization of homodimer/heterodimer on the cell membrane surface[38]; (B) HCR-based multi-F?rster Resonance Energy Transfer (multi-FRET) for imaging of cell membrane receptor glycosylation[40]; (C) FRET fluorescence signal switch and competitive fluorescence erasure strategy for imaging of cell membrane receptor glycosylation[41]

Figure 3. Structurally constrained hybridization probes for membrane receptor recognition in cell type identification (A) A pH-responsive activatable probe (pH-APP) for cell membrane receptor recognition[47]; (B) Cascade multiple structurally constrained fluorescence signal probes for highly sensitive imaging of cell membrane receptor dimerization[48]

Figure 4. Signal tag-based recognition of membrane receptors for cell type identification (A) A method for imaging cell membrane proteins by labeling SNAP fusion proteins in the live cell plasma membrane through the introduction of a charged sulfonate group (“SBG”) on the SNAP-tag substrate benzylguanine (BG)[62]; (B) A photocleavable peptide-labeled mass probe for imaging and mass spectrometry quantification of the cell membrane receptor HER2, achieved through probe binding, photocleavage released reporter peptide[66]

Figure 5. Imaging of membrane receptor interactions (A) A two-faced interactive DNA probe system for simultaneous monitoring of HER2/HER2 homodimerization and HER2/HER3 heterodimerization on the surface of tumor cells[77]; (B) A Nile Blue (NB)-conjugated sunitinib derivative biosensor selectively inserts into the RTKs domain pockets on the cancer cell membrane for imaging VEGFR-2 dimerization[78]; (C) A conformationally induced "off-on" biosensor, composed of sunitinib and pyrene, that recovers fluorescence in the presence of receptor protein tyrosine kinases, for imaging VEGFR-2 dimerization[79]; (D) scVEGF-conjugated fluorescent nanodiamonds (FND) target VEGFR to achieve imaging of membrane receptor-ligand interactions[80]

Figure 6. Imaging of membrane receptor spatial distribution (A) Aptamer-based live-cell PAINT technique for nanoscale imaging of membrane receptor spatial distribution through transient binding of aptamers to targets[86]; (B) Super-resolution microscopy combined with single-molecule FRET for the identification and imaging of EGF-activated ErbB1 dimers in living cells, enabling real-time monitoring of interactions between receptor proteins[87]; (C) The PD-PAINT method, which combines single-molecule localization super-resolution microscopy (SMLM) technology with a proximity-dependent DNA probe system, is used for detecting and imaging the spatial distribution of protein pairs in living cells[88]; (D) Surface plasmon-coupled emission (SPCE) strategy for spatial distribution imaging of cell membrane proteins by enhancing single-molecule blinking fluorescence through the fabrication of SPCE substrates[89]

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基于非基因工程的细胞膜受体蛋白可视化成像

Visualization Imaging of Cell Membrane Receptor Proteins Based on Non-genetic Engineering Approaches

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