Home Journals Articles Updates About
CN
image
收藏切换
Utilizing bivalent aptamers as first DNA agonist to activate RTKs heterodimer of different families
收藏切换
PDF
Kun Liu1, Yulin Cong1, Xiongfeng Luo, Meicun Yao*, Zhiyong Xie*, Hao Li*
Chinese Chemical Letters | 2025, 36(1) : 109839
Less
收藏切换
Chinese Chemical Letters | 2025, 36(1): 109839
• Communication •
Utilizing bivalent aptamers as first DNA agonist to activate RTKs heterodimer of different families
Full
Kun Liu1, Yulin Cong1, Xiongfeng Luo, Meicun Yao*, Zhiyong Xie*, Hao Li*
Affiliations
  • School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Shenzhen 518107, China
Published: 2025-01-15 doi: 10.1016/j.cclet.2024.109839
Outline
收藏切换

Heterodimerization in RTKs is of vital importance in the RTK signaling and cell functions. Heterodimerization between RTKs can result in diversity of downstream signals, increasing the ability of cells to respond to external experiments. Traditional RTKs heterodimerization always occur in the same families and is lack of agonists to activate the heterodimeric RTKs signaling pathway. Herein, we developed the DNA agonist based on bivalent aptamers for the heterodimerized RTKs of different families, AF/AM-1, which could simultaneously activate FGFR1 and c-Met signaling. It is the first agonist that realizing the heterodimerization and activation of FGFR1 and c-Met, two different RTK families. The activation of FGFR1/c-Met heterodimer result in the down-stream signals transduction, such as the phosphorylation of Akt and Erk, inducing the cell migration and proliferation. The DNA agonist for RTK heterodimer of different families would have potential applications in the fields of biomedicine.

DNA Agonist  /  Bivalent aptamers  /  RTKs heterodimer
Kun Liu, Yulin Cong, Xiongfeng Luo, Meicun Yao, Zhiyong Xie, Hao Li. Utilizing bivalent aptamers as first DNA agonist to activate RTKs heterodimer of different families[J]. Chinese Chemical Letters, 2025 , 36 (1) : 109839 - . DOI: 10.1016/j.cclet.2024.109839
Full Text
Less
Receptor tyrosine kinase, known as RTKs, are of great importance in cell survival and proliferation as well as differentiation [1,2]. Conventional activation of RTKs is based on the homodimerization between receptors through the binding of ligands which always be called agonists, such as various of growth factors [3]. However, the RTKs heterodimer remains insufficient agonists.
Heterodimerization is a common phenomenon among RTK receptors, and the cross-activation of signal pathways often occurs through heterodimerization [4,5]. For example, in hematopoietic cells, activation of the ErbB-3/ErbB-2 heterodimeric receptor triggers the formation and diffusion of PI3 kinase-dependent lamellipodia [6]. The degree of heterodimerization between RTK receptors depends on the concentration of receptors and ligands, which is not specific. In the ErbB family, EGF promotes the homodimerization of EGFR, but in fact, EGF has a higher affinity for the EGFR/ErbB2 heterodimer [7]. Heterodimerization between RTK receptors can alter their signal strength and duration, increase the diversity of downstream signals, and thereby increase the ability of cells to respond to external changes and stimuli [8-10]. Therefore, the development of RTK heterodimerization agonists has important implications in biomedical fields. However, most heterodimeric RTK receptors have insufficient corresponding agonists and the heterodimerization always occur between same RTK families. There is an urgent need for developing agonist of heterodimeric RTK receptors. Moreover, the realization of RTKs heterodimer between different families is waiting for new agonists. Currently, Inooka has successfully developed an artificial peptide agonist for the FGFR1c/KLB heterodimeric complex [11]. However, the preparation process of this dimeric peptide is complex and expensive. Moreover, peptides are not convenient to store and are easily degraded by proteases at the area of the wound. Therefore, developing novel heterodimeric receptor agonists is urgently needed.
Deoxyribonucleic acids (DNAs) are widely used in the fields of biosensing and cell regulation owing to their predictable hybridization and programmable assembly [12-17]. Functional DNAs, such as aptamers, could bind to the target with high specificity and affinity [18-23]. Based on the property of aptamers specifically binding to cell receptors, we and others have been developed using aptamers to construct molecular devices that activate cell receptor signal pathways [24-31]. We propose to use aptamers to construct novel DNA agonists for the activation of heterodimeric RTK receptors.
Herein, we developed a universal method to construct novel DNA agonist for the activation of heterodimeric RTK receptors between different families based on the bivalent aptamers. FGFR1 and c-Met, two different RTKs for fibroblast growth factor and hepatic growth factor respectively, were used as a model. FGFR1 and c-Met both could result in the activation of Akt and Erk signaling pathways, playing important role in cell migration and proliferation [24-31]. The research on simultaneously activation of FGFR1 and c-Met pathways is of great significance. Interestingly, the DNA agonist reported in current work realized the simultaneously activation of c-Met and FGFR1 based on the heterodimerization between c-Met and FGFR1. Moreover, the DNA agonist could result in the simultaneously activation of c-Met and EGFR [32], which shows good universality. We believe that the proposed DNA agonist will have broad applicability in cell therapy and regenerative medicine.
Our DNA agonist is composed of two subunits, AM and AF, hooking on the extracellular region of c-Met and FGFR1 receptor via the specific aptamer, respectively (Fig. 1). AM and AF could form a heterodimer through hybridization (Fig. 1 and Fig. S1 and Table S1 in Supporting information). The hybridization by AM and AF result in the heterodimerization of c-Met and FGFR1, following by the activation of the c-Met and FGFR1 signal pathway, which could induce the cell migration and proliferation.
We have developed a method that assembling AF/AM-R on the cell membrane (Fig. 2a Figs. S2 and S3 in Supporting information). As shown in Fig. 2a, the strands AF and AMR-TAMRA (AMRT) were both blocked by a complementary strand, AFRC and AMR-BHQ2 (AMRB), respectively, called AFR and AMR. The strands AF and AM could hook on the FGFR1 and c-Met [22,29]. In the presence of an initiate strand (AFRS), the AFR and AMR could interaction with each other, which could form the agonist AF/AM and bring the FGFR1 and c-Met heterodimerization. As shown in Fig. 2b, with the formation of AF/AM, the quenched TAMRA fluorescence significantly recovered. These results indicated that the agonist AF/AM could successfully assembling and hooking on the c-Met and FGFR1. The results were also demonstrated by turn-off confocal and flow cytometry experiments (Figs. S4 and S5 in Supporting information). The agonist, AF/AM-1, would bring anchored c-Met and FGFR1 in close proximity for cross phosphorylation. Indeed, AF/AM-1 quickly increased the c-Met and FGFR1 phosphorylation levels of A549 cells in 5 min (Fig. S6 in Supporting information). We have investigated the results about simply mixing FGFR1 and c-Met aptamers without complementary sequence. As shown in Fig. S7 (Supporting information), AF/AM-1 exhibit the highest c-Met and FGFR1 phosphorylation level, compared to the control that simply mixing FGFR1 and c-Met aptamers without complementary sequence. The length of complementary bases plays important role in phosphorylation of c-Met and FGFR1. As shown in Fig. S8 (Supporting information), the samples of different length, 12, 16, 23 (AF/AM-1), 30 and 40 were investigated. The 23 bp (AF/AM-1) length of complementary bases exhibited the highest phosphorylation level of FGFR1 and c-Met. Moreover, AF/AM-1 resulted in the FGFR1 and c-Met activation in a dose-dependent manner, with an effective dose as low as 10 nmol/L (Figs. 2c and d, Figs. S9 and S10 in Supporting information). Together, we have developed and characterized a novel heterodimerization agonist AF/AM-1 simultaneously activating FGFR1 and c-Met.
Next, we evaluated the downstream activation of Akt and Erk, key regulators in cell motility and cell proliferation (Fig. 3a). The A549 cells responded to AF/AM-1 with increased phosphorylation of both Akt and Erk in a linear manner (Fig. 3b, Figs. S11 and S12 in Supporting information). In response to AF/AM-1, GFP-actin transfected A549 cells generated large amounts of the actin lamellipodium (Fig. 3c), which indicate the potential increased motility, whereas control cells did not respond. The proliferation of AF/AM-1 treated A549 cells was 1.2x than untreated cells in 24 h (Fig. 3d). As expected, AF/AM-1 improved the cell motility compared with untreated cells (Fig. 3e). The AF/AM-1 treated cell motility rate was 16.4 %, which was 3.1x compared with the control group (Fig. 3f). Above all, we have demonstrated the DNA agonist AF/AM-1 could induce the phosphorylation of both Akt and Erk, which successfully result in the enhanced cell motility and proliferation.
To validate the DNA agonist system universality for application, we designed three different DNA agonists, AF-AM, AF/AM-1 and AF/AM-2 (Fig. 4a and Fig. S1). All the three agonists exhibited distinctly phosphorylation of c-Met and FGFR1 (Fig. 4b). As shown in Figs. 4c and d, the AF/AM-1 exhibited the best phosphorylation level of c-Met and FGFR1. It is mainly due to the distance of two aptamers in the agonist AF/AM-1, compared with the AF-AM and AF/AM-2. In AF/AM-1, the distance about c-Met and FGFR1 aptamer is 23 bp, however, in AF-AM and AF/AM-2, the distance is 0 bp, as shown in Fig. 4a. The distance between two aptamers is of great importance in the activation of c-Met and FGFR1, as the results in Fig. S8 (Supporting information). The distance between c-Met and FGFR1 aptamer in AF/AM-1 may be more suit to anchor c-Met and FGFR1 simultaneously, resulting in the best phosphorylation of c-Met and FGFR1. Moreover, all the three agonists resulted in the obviously proliferation of A549 cells (Fig. S13 in Supporting information). Then, we evaluated the feasibility of our agonists in the other cells (Figs. S14–S18 in Supporting information). These results demonstrated that the design of our DNA agonists exhibited good generality. Next, we evaluate the feasibility of our agonists in the other heterodimerization receptor pairs, for instance, EGFR/c-Met receptor heterodimerization and EGFR/FGFR1 (Fig. 4e and Fig. S20 in Supporting information). The two agonists AE/AM-1 and AE/AM-2 both exhibited the enhanced phosphorylation level of EGFR and c-Met (Figs. 4f–h). As shown in Fig. S19 (Supporting information), simply mixing EGFR and c-Met aptamers without complementary sequence could hardly activate EGFR and c-Met phosphorylation. In addition, the two agonists AF/AE-1 and AF/AE-2 both exhibited the enhanced phosphorylation level of FGFR1 and EGFR (Fig. S20).
In summary, we have developed a universal method to construct DNA agonist for the activation of the heterodimerization RTK receptors between different families. To the best of our knowledge, this is the first example of using DNA agonist for the activation of heterodimerization RTK receptors and signaling pathways. The DNA agonist shows excellent versatility in potential applications owing to (1) the flexibility of the recognition module of the agonist, which may include various aptamers binding for different receptors, allowing for a wide range of receptor signaling pathways to be used as specific applications, and (2) the customizability of the linking modules with other hybridization manner or strand displacement reaction approach, establishing the possibility to expand available DNA nanotechnology and DNA nanodevices. Notably, we have simultaneously controlled target receptors heterodimerization and activated the receptor signaling pathways by using DNA agonists, potentially offering a facile and practical nongenetic approach for customized cell behaviors in vivo by receptors heterodimerization in the future. This strategy represents a new avenue for the smart regulation of cell behavior in cell-based therapy and regenerative medicine.
Declaration of competing interest
Less
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
CRediT authorship contribution statement
Less
Kun Liu: Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Validation, Visualization, Writing – original draft, Writing – review & editing. Yulin Cong: Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Validation, Visualization, Writing – original draft, Writing – review & editing. Xiongfeng Luo: Data curation, Formal analysis, Investigation, Software, Validation, Visualization. Meicun Yao: Conceptualization, Project administration, Resources, Software, Supervision, Writing – review & editing. Zhiyong Xie: Conceptualization, Funding acquisition, Resources, Software, Supervision, Validation, Writing – review & editing. Hao Li: Conceptualization, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Validation, Visualization, Writing – original draft, Writing – review & editing.
Acknowledgments
Less
The authors are grateful to the National Natural Science Foundation of China (Nos. 22104158, 82174104, U1903211), the Natural Science Foundation of Hunan Province (No. 2021JJ40041), Guangdong Basic and Applied Basic Research Foundation (No. 2023A1515011346), Shenzhen Science and Technology Program (No. SZBH202130).
Supplementary materials
Less
Supplementary material associated with this article can be found, in the online version, at doi:10.1016/j.cclet.2024.109839.
References
Less
[1]
J. Li, L. Wang, J. Tian, et al., Chem. Soc. Rev. 49 (2020) 1545–1568.
[2]
Z. Du, C.M. Lovly, Mol. Cancer 17 (2018) 58.
[3]
M.A. Lemmon, J. Schlessinger, Cell 141 (2010) 1117–1134.
[4]
L. Chen, L. Fu, J. Sun, et al., Nature 618 (2023) 862–870.
[5]
M.D. Paul, K. Hristova, Chem. Rev. 119 (2019) 5881–5921.
[6]
A. Chausovsky, H. Waterman, M. Elbaum, et al., Oncogene 19 (2000) 878–888.
[7]
J.L. Macdonald-Obermann, L.J. Pike, J. Biol. Chem. 289 (2014) 26178–26188.
[8]
M. Landau, N. Ben-Tal, Biochim. Biophys. Acta 1785 (2008) 12–31.
[9]
M.A. Olayioye, R.M. Neve, H.A. Lane, et al., EMBO J. 19 (2000) 3159–3167.
[10]
Y. Yarden, M.X. Sliwkowski, Nat. Rev. Mol. Cell. Biol. 2 (2001) 127–137.
[11]
K. Sakamoto, Y. Kawata, Y. Masuda, et al., Biochem. Biophys. Res. Commun. 480 (2016) 55–60.
[12]
Y. Du, Y. Lyu, J. Lin, et al., Nat. Nanotechnol. 18 (2023) 818–827.
[13]
D.Y. Zhang, A.J. Turberfield, B. Yurke, et al., Science 318 (2007) 1121–1125.
[14]
X. Mao, M. Liu, Q. Li, et al., JACS Au 2 (2022) 2381–2399.
[15]
Y.J. Chen, B. Groves, R.A. Muscat, et al., Nat. Nanotechnol. 10 (2015) 748–760.
[16]
Y. Tang, Z. Wang, X. Yang, et al., Chem. Sci. 6 (2015) 5729–5733.
[17]
M. You, L. Peng, N. Shao, et al., J. Am. Chem. Soc. 136 (2014) 1256–1259.
[18]
R. Nutiu, Y. Li, J. Am. Chem. Soc. 125 (2003) 4771–4778.
[19]
J. Liu, Z. Cao, Y. Lu, Chem. Rev. 109 (2009) 1948–1998.
[20]
N. Nakatsuka, K.A. Yang, J.M. Abendroth, et al., Science 362 (2018) 319–324.
[21]
W. Tan, M.J. Donovan, J. Jiang, Chem. Rev. 113 (2013) 2842–2862.
[22]
K. Zhang, H. Gao, R. Deng, et al., Angew. Chem. Int. Ed. 58 (2019) 4790–4799.
[23]
K. Zhang, R. Deng, Y. Sun, et al., Chem. Sci. 8 (2017) 7098–7105.
[24]
H. Li, M. Wang, T. Shi, et al., Angew. Chem. Int. Ed. 57 (2018) 10226–10230.
[25]
H. Li, J. Gao, L. Cao, et al., Angew. Chem. Int. Ed. 60 (2021) 26087–26095.
[26]
R. Ueki, A. Ueki, N. Kanda, et al., Angew. Chem. Int. Ed. 55 (2016) 579–582.
[27]
H. Liang, Z. Yan, Y. Tong, et al., Chem. Commun. 59 (2023) 1621–1624.
[28]
M. Wang, F. He, H. Li, et al., Nano. Lett. 19 (2019) 2603–2613.
[29]
L. Wang, H. Liang, J. Sun, et al., J. Am. Chem. Soc. 141 (2019) 12673–12681.
[30]
R. Ueki, S. Sando, Chem. Commun. 50 (2014) 13131–13134.
[31]
R. Ueki, S. Atsuta, A. Ueki, et al., Chem. Commun. 55 (2019) 2672–2675.
[32]
D.L. Wang, Y.L. Song, Z. Zhu, et al., Biochem. Biophys. Res. Commun. 453 (2014) 681–685.
Appendix
Less
Year 2025 volume 36 Issue 1
PDF
58
31
Cite this Article
BibTeX
Article Info
doi: 10.1016/j.cclet.2024.109839
  • Receive Date：2024-02-04
  • Online Date：2025-11-12
  • Published：2025-01-15
Article Data
Affiliations
History
  • Received：2024-02-04
  • Revised：2024-03-01
  • Accepted：2024-03-27
Affiliations
    School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Shenzhen 518107, China
References
Share
https://castjournals.cast.org.cn/joweb/ccl/EN/10.1016/j.cclet.2024.109839
Share to
QR

Scan QR to access full text

Cite this article
BibTeX
Citations
表12种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
Genus		 种数
Number of
species		 占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
关闭全屏
  • BibTeX
  • EndNote
  • RefWorks
  • TxT
Links
Articles
Latest Articles
Most Read
Collections
Updates
Events
News
Multimedia
About
About Us
Contact
No. 86 Xueyuan South Road, Haidian District, Beijing
100081
010-62199257
qkjq@cast.org.cn

Copyright © 2025 China Association for Science and Technology. All rights reserved. For all open access content, the relevant licensing terms apply.
Sponsored by the Office of the Leading Group for Cybersecurity and Informatization of CAST, and supported by Science and Technology Review Publishing House