Establishment of a model of developmentally regulated brain protein 1 knockdown based on the Tet-on system and its regulatory effect on nuclear morphological homeostasis and senescence of lung cancer cells

Hu Xing; Li Yao; Zhang Lian; Zhang Chundong

doi:10.13406/j.cnki.cyxb.004092

您当前的位置：

首页 >

文章列表页 >

Establishment of a model of developmentally regulated brain protein 1 knockdown based on the Tet-on system and its regulatory effect on nuclear morphological homeostasis and senescence of lung cancer cells

Basic research | 更新时间：2026-05-13

- Establishment of a model of developmentally regulated brain protein 1 knockdown based on the Tet-on system and its regulatory effect on nuclear morphological homeostasis and senescence of lung cancer cells
- Journal of Chongqing Medical University Vol. 51, Issue 5, Pages: 645-653(2026)
- 作者机构：
  
  1.重庆医科大学生物化学与分子生物学教研室，重庆 400016
  2.重庆医科大学组织学与胚胎学教研室，重庆 400016
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.004092
  CLC： R734.2
- Received：16 January 2026，
  
  Revised：2026-03-19，
  
  Accepted：20 March 2026，
  
  Published：28 May 2026
- 稿件说明：
移动端阅览
胡星，李瑶，张莲，等.基于Tet-on系统的DBN1诱导敲减模型的构建及其对肺癌细胞核形态稳态与衰老的调控作用[J].重庆医科大学学报，2026，51（5）：645-653.

Hu Xing,Li Yao,Zhang Lian,et al.Establishment of a model of developmentally regulated brain protein 1 knockdown based on the Tet-on system and its regulatory effect on nuclear morphological homeostasis and senescence of lung cancer cells[J].Journal of Chongqing Medical University,2026,51(5):645-653.
胡星，李瑶，张莲，等.基于Tet-on系统的DBN1诱导敲减模型的构建及其对肺癌细胞核形态稳态与衰老的调控作用[J].重庆医科大学学报，2026，51（5）：645-653. DOI： 10.13406/j.cnki.cyxb.004092.

Hu Xing,Li Yao,Zhang Lian,et al.Establishment of a model of developmentally regulated brain protein 1 knockdown based on the Tet-on system and its regulatory effect on nuclear morphological homeostasis and senescence of lung cancer cells[J].Journal of Chongqing Medical University,2026,51(5):645-653. DOI： 10.13406/j.cnki.cyxb.004092.

摘要

目的

利用Tet-on慢病毒诱导系统构建发展调节脑蛋白1（developmentally regulated brain protein 1，DBN1）（Drebrin）条件性敲减的肺癌A549细胞模型，探讨DBN1在肺癌恶性表型维持、细胞核形态稳态及细胞衰老中的作用。

方法

通过UALCAN和基因表达谱交互分析数据库（Gene Expression Profiling Interactive Analysis database，GEPIA）分析DBN1在肺癌中的表达特征及预后价值；构建pInducer10-shDBN1诱导表达系统，筛选并优化Doxy诱导条件；采用蛋白质印迹（Western blot）、克隆形成、划痕实验及β-半乳糖苷酶染色等方法，系统评估DBN1缺失对细胞增殖、迁移、衰老及细胞核形态的影响。

结果

患者预后不良与肺癌组织中DBN1的异常上调显著相关，临床数据显示该蛋白在癌组织中呈现高丰度表达（

0.050）。成功构建高效、可控的Tet-on诱导敲减模型（4 μg/mL Doxy诱导6 d为最佳条件）。功能实验表明，诱导敲减DBN1明显抑

制A549细胞的增殖与迁移（

0.010），并诱发明显的细胞衰老表型［衰老相关分泌表型因子（senescence-associated secretory phenotype factor，SASP）白细胞介素-6（interleukin-6，IL-6）、肿瘤坏死因子（tumor necrosis factor，TNF）等上调］。免疫荧光显示，DBN1缺失导致F-actin重构异常及核纤层蛋白Lamin A/C分布紊乱，引起细胞核形态扭曲。

结论

DBN1是肺癌进展的关键促癌因子；本研究通过诱导系统揭示DBN1在维持肺癌细胞核形态稳态及抑制衰老中的新功能，为肺癌靶向“诱导衰老”治疗提供潜在靶点。

Abstract

Objective

To establish a conditional knockdown model of developmentally regulated brain protein 1（DBN1，also known as Drebrin） in human lung cancer A549 cells using the inducible Tet‑on lentivirus system，and to investigate the role of DBN1 in maintaining malignant phenotype，preserving nuclear morphological homeostasis，and regulating cellular senescence in lung cancer.

Methods

UALCAN and Gene Expression Profiling Interactive Analysis database were used to analyze the expression characteristics and prognostic value of DBN1 in lung cancer. The pInducer10-shDBN1 inducible expression system was constructed，and doxycycline induction conditions were screened and optimized. Western blot，colony formation assay，wound healing assay，and β-galactosidase staining were used to assess the effect of DBN1 depletion on cell proliferation，migration，senescence，and nuclear morphology.

Results

The poor prognosis of patients was significantly associated with abnormal upregulation of DBN1 in lung cancer tissue，and clinical data showed that this protein was highly abundant in cancerous tissue（

0.05）. An efficient and controllable Tet‑on inducible knockdown model was successfully established，with the optimal condition of induction with 4 μg/mL doxycycline for 6 days. Functional experiments showed that inducible knockdown of DBN1 significantly inhibited the proliferation and migration of A549 cells（

0.01） and triggered a pronounced cellular senescence phenotype，accompanied by upregulation of the

senescence‑associated secretory phenotype factors such as interleukin‑6 and tumor necrosis factor. Immunofluorescence assay showed that DBN1 depletion resulted in abnormal F‑actin remodeling and disrupted distribution of the nuclear lamina proteins Lamin A/C，thereby leading to distorted nuclear morphology.

Conclusion

DBN1 is a key oncogenic factor that promotes the progression of lung cancer. By using an inducible knockdown system，this study reveals a novel role of DBN1 in maintaining nuclear morphological homeostasis and inhibiting cellular senescence in lung cancer and provides a potential therapeutic target for senescence-inducing treatment of lung cancer.

关键词

Keywords

references

Bray F ， Laversanne M ， Sung H ， et al . Global cancer statistics 2022： GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries [J ] . CA Cancer J Clin ， 2024 ， 74 （ 3 ）： 229 - 263 .

Han BF ， Zheng RS ， Zeng HM ， et al . Cancer incidence and mortality in China，2022 [J ] . J Natl Cancer Cent ， 2024 ， 4 （ 1 ）： 47 - 53 ．

中华医学会肿瘤学分会，韩宝惠，王　洁，等 . 中华医学会肺癌临床诊疗指南（2025版） [J ] . 中华肿瘤杂志， 2025 ， 47 （ 9 ）： 769 - 810 ．

Oncology Society of Chinese Medical Association ， Han BH ， Wang J ， et al . Chinese Medical Association guideline for clinical diagnosis and treatment of lung cancer（2025 edition） [J ] . Chin J Oncol ， 2025 ， 47 （ 9 ）： 769 - 810 ．

Wu FY ， Fan J ， He YY ， et al . Single-cell profiling of tumor heterogeneity and the microenvironment in advanced non-small cell lung cancer [J ] . Nat Commun ， 2021 ， 12 （ 1 ）： 2540 ．

Shirao T ， Obata K . Two acidic proteins associated with brain development in chick embryo [J ] . J Neurochem ， 1985 ， 44 （ 4 ）： 1210 - 1216 ．

Kojima N ， Shirao T ， Obata K . Molecular cloning of a developmentally regulated brain protein，chicken drebrin A and its expression by alternative splicing of the drebrin gene [J ] . Brain Res Mol Brain Res ， 1993 ， 19 （ 1/2 ）： 101 - 114 ．

Shirao T ， Sekino Y . Drebrin：From Structure and Function to Physiological and Pathological Roles [M ] . Tokyo ： Springer Japan ， 2017 ．

Ishikawa R ， Hayashi K ， Shirao T ， et al . Drebrin，a development-associated brain protein from rat embryo，causes the dissociation of tropomyosin from actin filaments [J ] . J Biol Chem ， 1994 ， 269 （ 47 ）： 29928 - 29933 ．

Dart AE ， Gordon-Weeks PR . The Role of Drebrin in Cancer Cell Invasion ， in Drebrin : From Structure and Function to Physiological and Pathological Roles ， T . Shirao and Y. Sekino ，Editors. 2017 ，Springer Japan: Tokyo. p. 375 - 389 .

Monami G ， Emiliozzi V ， Bitto A ， et al . Proepithelin regulates prostate cancer cell biology by promoting cell growth，migration，and anchorage-independent growth [J ] . Am J Pathol ， 2009 ， 174 （ 3 ）： 1037 - 1047 .

Terakawa Y ， Agnihotri S ， Golbourn B ， et al . The role of drebrin in glioma migration and invasion [J ] . Exp Cell Res ， 2013 ， 319 （ 4 ）： 517 - 528 .

Sun Y ， Wei HN ， Yu WT ， et al . The actin-binding protein drebrin disrupts NF2-LATS kinases complex assembly to facilitate liver tumorigenesis [J ] . Hepatology ， 2025 ， 81 （ 5 ）： 1433 - 1451 ．

Iyama S ， Ono M ， Kawai-Nakahara H ， et al . Drebrin： A new oncofetal biomarker associated with prognosis of lung adenocarcinoma [J ] . Lung Cancer ， 2016 ， 102 ： 74 - 81 ．

Yu QY ， Tang FY ， Cao FY ， et al . Drebrin promotes lung adenocarcinoma cell migration through inducing integrin β1 endocytosis [J ] . Biochem Biophys Res Commun ， 2022 ， 630 ： 175 - 182 ．

Meerbrey KL ， Hu G ， Kessler JD ， et al . The pINDUCER lentiviral toolkit for inducible RNA interference in vitro and in vivo [J ] . Proc Natl Acad Sci USA ， 2011 ， 108 （ 9 ）： 3665 - 3670 ．

Li Y ， Song MY ， Hu X ， et al . TNIK regulates cytoskeletal organization to promote focal adhesion turnover and mitosis in lung adenocarcinoma [J ] . Front Biosci ， 2025 ， 30 （ 5 ）： 38875 ．

Shirao T ， Sekino Y . Drebrin：From Structure and Function to Physiological and Pathological Roles [M ] . Tokyo ： Springer Japan ， 2017 ．

Mishra S ， Levy DL . Nuclear F-actin and Lamin A antagonistically modulate nuclear shape [J ] . J Cell Sci ， 2022 ， 135 （ 13 ）： jcs259692 ．

Kalukula Y ， Stephens AD ， Lammerding J ， et al . Mechanics and functional consequences of nuclear deformations [J ] . Nat Rev Mol Cell Biol ， 2022 ， 23 （ 9 ）： 583 - 602 ．

Vahabikashi A ， Sivagurunathan S ， Nicdao FAS ， et al . Nuclear lamin isoforms differentially contribute to LINC complex-dependent nucleocytoskeletal coupling and whole-cell mechanics [J ] . Proc Natl Acad Sci USA ， 2022 ， 119 （ 17 ）： e2121816119 ．

Wang X ， Björklund S ， Wasik AM ， et al . Gene expression profiling and chromatin immunoprecipitation identify DBN1 SETMAR and HIG2 as direct targets of SOX11 in mantle cell lymphoma [J ] . PLoS One ， 2010 ， 5 （ 11 ）： e14085 ．

Keon BH ， Jedrzejewski PT ， Paul DL ， et al . Isoform specific expression of the neuronal F-actin binding protein，drebrin，in specialized cells of stomach and kidney epithelia [J ] . J Cell Sci ， 2000 ， 113 Pt 2 ： 325 - 336 ．

Peitsch WK ， Grund C ， Kuhn C ， et al . Drebrin is a widespread actin-associating protein enriched at junctional plaques，defining a specific microfilament anchorage system in polar epithelial cells [J ] . Eur J Cell Biol ， 1999 ， 78 （ 11 ）： 767 - 778 ．

Peitsch WK ， Hofmann I ， Bulkescher J ， et al . Drebrin，an actin-binding，cell-type characteristic protein： induction and localization in epithelial skin tumors and cultured keratinocytes [J ] . J Invest Dermatol ， 2005 ， 125 （ 4 ）： 761 - 774 ．

Yagoub D ， Wilkins MR ， Lay AJ ， et al . Sphingosine kinase 1 isoform-specific interactions in breast cancer [J ] . Mol Endocrinol ， 2014 ， 28 （ 11 ）： 1899 - 1915 ．

Xu SQ ， Buraschi S ， Morcavallo A ， et al . A novel role for drebrin in regulating progranulin bioactivity in bladder cancer [J ] . Oncotarget ， 2015 ， 6 （ 13 ）： 10825 - 10839 ．

de Magalhães JP . Cellular senescence in normal physiology [J ] . Science ， 2024 ， 384 （ 6702 ）： 1300 - 1301 ．

Pathak RU ， Soujanya M ， Mishra RK . Deterioration of nuclear morphology and architecture： A hallmark of senescence and aging [J ] . Ageing Res Rev ， 2021 ， 67 ： 101264 ．

Colucci M ， Sarill M ， Maddalena M ， et al . Senescence in cancer [J ] . Cancer Cell ， 2025 ， 43 （ 7 ）： 1204 - 1226 ．

Sohrabi Molina M ， Brauer E ， Günther R ， et al . Increased substrate stiffness disrupts nuclear-cytoskeletal mechanical coupling in senescent cells [J ] . Mater Today Bio ， 2025 ， 35 ： 102472 ．

Zhou ZY ， Qin Q ， Dong CY ， et al . The dual guardians of cellular stability：exploring nesprin and lamin in senescence [J ] . Cell Death Dis ， 2025 ， 16 （ 1 ）： 757 ．

Dauban L ， Eder M ， de Haas M ， et al . Interactions between the genome and the nuclear Lamina are multivalent and cooperative [J ] . Nat Struct Mol Biol ， 2025 ， 32 （ 11 ）： 2335 - 2348 ．

Gholamalamdari O ， van Schaik T ， Wang YC ， et al . Major nuclear locales define nuclear genome organization and function beyond A and B compartments [J ] . Elife ， 2025 ，13：RP99116．

Lee S ， Schmitt CA . The dynamic nature of senescence in cancer [J ] . Nat Cell Biol ， 2019 ， 21 （ 1 ）： 94 - 101 ．

Wang LQ ， Lankhorst L ， Bernards R . Exploiting senescence for the treatment of cancer [J ] . Nat Rev Cancer ， 2022 ， 22 （ 6 ）： 340 - 355 ．

McHugh D ， Durán I ， Gil J . Senescence as a therapeutic target in cancer and age-related diseases [J ] . Nat Rev Drug Discov ， 2025 ， 24 （ 1 ）： 57 - 71 ．

Aoki C ， Sekino Y ， Hanamura K ， et al . Drebrin A is a postsynaptic protein that localizes in vivo to the submembranous surface of dendritic sites forming excitatory synapses [J ] . J Comp Neurol ， 2005 ， 483 （ 4 ）： 383 - 402 ．

Peitsch WK ， Hofmann I ， Endlich N ， et al . Cell biological and biochemical characterization of drebrin complexes in mesangial cells and podocytes of renal glomeruli [J ] . J Am Soc Nephrol ， 2003 ， 14 （ 6 ）： 1452 - 1463 ．

Views

下载量

CSCD

Alert me when the article has been cited

提交

Tools

Publicity Resources

Potential mechanisms and prospects of yin-nourishing traditional Chinese medicines in preventing and treating lung cancer based on aquaporins

Classification of the development trajectory of symptom distress and core influencing factors in patients with advanced lung cancer undergoing concurrent chemoradiotherapy based on the latent variable mixed growth model

Construction and validation of a risk prediction model for delayed chemotherapy-induced nausea and vomiting in patients with lung cancer

Effect of ATM/CHK2 regulation by miR-181a-5p on the senescence of human skin fibroblasts and its mechanism

Association between dynamic platelet count trajectories and adverse outcomes in patients with sepsis-induced coagulopathy

Related Author

Han Weiwei

Nie Peng

Chen Limei

Yan Xiaojun

Liu Hongning

Shan Si

Zou Xia

Li Haixiang

Related Institution

Affiliated Hospital of Jiangxi University of Chinese Medicine

Research Center for Differentiation and Development of TCM Basic Theory of Jiangxi University of Chinese Medicine，Jiangxi Province Key Laboratory of TCM Etiopathogenesis

Department of Oncology，Guangdong Second Provincial General Hospital，Jinan University

Department of Respiratory Medicine，The Affiliated Cancer Hospital of Zhengzhou University/ Henan Cancer Hospital

Department of Plastic and Burn Surgery，The First Affiliated Hospital of Chongqing Medical University

⁰