ACTN1 interacts with ITGA5 to promote cell proliferation, invasion and epithelial-mesenchymal transformation in head and neck squamous cell carcinoma

Document Type : Original Article


1 Department of Otolaryngology-Head and Neck Surgery, Shanxi Bethune Hospital, Shanxi Academy of Medical Sciences, Tongji Shanxi Hospital, Third Hospital of Shanxi Medical University, Taiyuan, Shanxi, 030032, China

2 Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China


Objective(s): The aim of this study was to detect the expression levels of α-Actinin 1 (ACTN1) and ITGA5 in HNSCC and to explore how ACTN1/ITGA5 regulated the proliferative and invasive abilities, as well as the EMT of Head and neck squamous cell carcinoma (HNSCC) cells. 
Materials and Methods: The viability, proliferative, invasive and migrative abilities of HNSCC cells after transfection were, in turn, detected by CCK‑8 assay, colony formation assay, EdU staining, transwell, as well as wound healing. E-cadherin in transfected cells was assessed utilizing immunofluorescence. RT-qPCR confirmed the transfection effect of ACTN1 and ITGA5 in HNSCC cells and the interaction between ACTN1 and ITGA5 in HNSCC cells was determined by co-immunoprecipitation (Co-IP). With Western blot application, the contents of ACTN1, ITGA5, proliferation-, invasion- and migration-related proteins were estimated. A xenograft model based on nude mice was conducted and Ki-67 content in tumor tissues was evaluated employing immunohistochemistry (IHC) staining.
Results: ACTN1 interacted with ITGA5. The contents of ACTN1 and ITGA5 were found to be abundant in HNSCC tissues and cells and associated with poor prognosis. ACTN1 depletion imparted suppressive impacts on cell proliferative, invasive and migrative abilities as well as EMT of HNSCC cells, which were reversed by ITGA5 overexpression. In addition, ACTN1 deficiency repressed the growth and metastasis of tumor tissues in tumor xenografts of nude mice. 
Conclusion: ACTN1 positively interacts with ITGA5 to promote proliferation, invasion and EMT of HNSCC cells. Also, ACTN1 promotes tumor growth and metastasis.


1. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 2018; 68: 394-424.
2.Chow LQM. Head and Neck Cancer. N Engl J Med 2020; 382: 60-72.
3. Wang Y, Wang S, Wu Y, Ren Y, Li Z, Yao X, et al. Suppression of the growth and invasion of human head and neck squamous cell carcinomas via regulating STAT3 signaling and the miR-21/β-catenin axis with HJC0152. Mol Cancer Ther 2017; 16: 578-590.
4. Lam L, Logan RM, Luke C. Epidemiological analysis of tongue cancer in South Australia for the 24-year period, 1977-2001. Aust Dent J 2006; 51: 16-22.
5. Puram SV, Rocco JW. Molecular aspects of head and neck cancer therapy. Hematol Oncol Clin North Am 2015; 29:971-992.
6. Dixson JD, Forstner MJ, Garcia DM. The alpha-actinin gene family: a revised classification. J Mol Evol 2003; 56: 1-10.
7. Parr T, Waites GT, Patel B, Millake DB, Critchley DR. A chick skeletal-muscle alpha-actinin gene gives rise to two alternatively spliced isoforms which differ in the EF-hand Ca(2+)-binding domain. Eur J Biochem 1992; 210: 801-809.
8. Wyszynski M, Kharazia V, Shanghvi R, Rao A, Beggs AH, Craig AM, et al. Differential regional expression and ultrastructural localization of alpha-actinin-2, a putative NMDA receptor-anchoring protein, in rat brain. J Neurosci 1998; 18: 1383-1392.
9. Craig DH, Downey C, Basson MD. SiRNA-mediated reduction of alpha-actinin-1 inhibits pressure-induced murine tumor cell wound implantation and enhances tumor-free survival. Neoplasia 2008; 10: 217-222.
10. Chen Q, Zhou XW, Zhang AJ, He K. ACTN1 supports tumor growth by inhibiting Hippo signaling in hepatocellular carcinoma. J Exp Clin Cancer Res 2021; 40: 23-35.
11. Cao Y, Cao W, Qiu Y, Zhou Y, Guo Q, Gao Y, et al. Oroxylin A suppresses ACTN1 expression to inactivate cancer-associated fibroblasts and restrain breast cancer metastasis. Pharmacol Res 2020; 159: 104981.
12. Xie GF, Zhao LD, Chen Q, Tang DX, Chen QY, Lu HF, et al. High ACTN1 is associated with poor prognosis, and ACTN1 silencing suppresses cell proliferation and metastasis in oral squamous cell carcinoma. Drug Des Devel Ther 2020; 14: 1717-1727.
13. Morgan MR, Byron A, Humphries MJ, Bass MD. Giving off mixed signals--distinct functions of alpha5beta1 and alphavbeta3 integrins in regulating cell behaviour. IUBMB Life 2009; 61: 731-738.
14. Feng C, Jin X, Han Y, Guo R, Zou J, Li Y, et al. Expression and prognostic analyses of ITGA3, ITGA5, and ITGA6 in head and neck squamous cell carcinoma. Med Sci Monit 2020; 26: e926800.
15. Pan L, Yang H, Xu C, Chen S, Meng Z, Li K, et al. ZNF750 inhibited the malignant progression of oral squamous cell carcinoma by regulating tumor vascular microenvironment. Biomed Pharmacother 2018; 105: 566-572.
16. Li T, Wu Q, Liu D, Wang X. miR-27b suppresses tongue squamous cell carcinoma epithelial-mesenchymal transition by targeting ITGA5. Onco Targets Ther 2020; 13: 11855-11867.
17. Deng Y, Wan Q, Yan W. Integrin α5/ITGA5 promotes the proliferation, migration, invasion and progression of oral squamous carcinoma by epithelial-mesenchymal transition. Cancer Manag Res 2019; 11: 9609-9620.
18. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 2001; 25: 402-408.
19. Marur S, Forastiere AA. Head and neck squamous cell carcinoma: update on epidemiology, diagnosis, and treatment. Mayo Clin Proc 2016; 91: 386-396.
20. Liu C, Yu Z, Huang S, Zhao Q, Sun Z, Fletcher C, et al. Combined identification of three miRNAs in serum as effective diagnostic biomarkers for HNSCC. EBioMedicine 2019; 50: 135-143.
21. Liu Z, Zhan Y, Tu Y, Chen K, Liu Z, Wu C. PDZ and LIM domain protein 1(PDLIM1)/CLP36 promotes breast cancer cell migration, invasion and metastasis through interaction with α-actinin. Oncogene 2015; 34: 1300-1311.
22. Ngan E, Northey JJ, Brown CM, Ursini-Siegel J, Siegel PM. A complex containing LPP and α-actinin mediates TGFβ-induced migration and invasion of ErbB2-expressing breast cancer cells. J Cell Sci 2013; 126: 1981-1991.
23. Honda K, Yamada T, Endo R, Ino Y, Gotoh M, Tsuda H, et al. Actinin-4, a novel actin-bundling protein associated with cell motility and cancer invasion. J Cell Biol 1998; 140: 1383-1393.
24. Honda K, Yamada T, Hayashida Y, Idogawa M, Sato S, Hasegawa F, et al. Actinin-4 increases cell motility and promotes lymph node metastasis of colorectal cancer. Gastroenterology 2005; 128: 51-62.
25. Kikuchi S, Honda K, Tsuda H, Hiraoka N, Imoto I, Kosuge T, et al. Expression and gene amplification of actinin-4 in invasive ductal carcinoma of the pancreas. Clin Cancer Res 2008; 14: 5348-5356.
26. Yamamoto S, Tsuda H, Honda K, Kita T, Takano M, Tamai S, et al. Actinin-4 expression in ovarian cancer: a novel prognostic indicator independent of clinical stage and histological type. Mod Pathol 2007; 20:1278-1285.
27. Barbolina MV, Adley BP, Kelly DL, Fought AJ, Scholtens DM, Shea LD, et al. Motility-related actinin alpha-4 is associated with advanced and metastatic ovarian carcinoma. Lab Invest 2008; 88: 602-614.
28. Xu S, Cui L, Ma D, Sun W, Wu B. Effect of ITGA5 down-regulation on the migration capacity of human dental pulp stem cells. Int J Clin Exp Pathol 2015; 8: 14425-14432.
29. Foubert P, Varner JA. Integrins in tumor angiogenesis and lymphangiogenesis. Methods Mol Biol 2012; 757: 471-486.
30. Wu D, Xu Y, Ding T, Zu Y, Yang C, Yu L. Pairing of integrins with ECM proteins determines migrasome formation. Cell Res 2017; 27: 1397-1400.
31. Das V, Kalyan G, Hazra S, Pal M. Understanding the role of structural integrity and differential expression of integrin profiling to identify potential therapeutic targets in breast cancer. J Cell Physiol 2018; 233: 168-185.
32. Cao L, Chen Y, Zhang M, Xu DQ, Liu Y, Liu T, et al. Identification of hub genes and potential molecular mechanisms in gastric cancer by integrated bioinformatics analysis. PeerJ 2018; 6: e5180.
33. Xiao Y, Li Y, Tao H, Humphries B, Li A, Jiang Y, et al. Integrin α5 down-regulation by miR-205 suppresses triple negative breast cancer stemness and metastasis by inhibiting the Src/Vav2/Rac1 pathway. Cancer Lett 2018; 433: 199-209.
34. Gao Q, Yang Z, Xu S, Li X, Yang X, Jin P, et al. Heterotypic CAF-tumor spheroids promote early peritoneal metastatis of ovarian cancer. J Exp Med 2019; 216: 688-703.
35. Chen J, Ji T, Wu D, Jiang S, Zhao J, Lin H, et al. Human mesenchymal stem cells promote tumor growth via MAPK pathway and metastasis by epithelial mesenchymal transition and integrin α5 in hepatocellular carcinoma. Cell Death Dis 2019; 10: 425-436.
36. Li XQ, Lu JT, Tan CC, Wang QS, Feng YM. RUNX2 promotes breast cancer bone metastasis by increasing integrin α5-mediated colonization. Cancer Lett 2016; 380: 78-86.
37. Zhang XB, Song L, Wen HJ, Bai XX, Li ZJ, Ma LJ. Upregulation of microRNA-31 targeting integrin α5 suppresses tumor cell invasion and metastasis by indirectly regulating PI3K/AKT pathway in human gastric cancer SGC7901 cells. Tumour Biol 2016; 37: 8317-8325.
38. Fan QC, Tian H, Wang Y, Liu XB. Integrin-α5 promoted the progression of oral squamous cell carcinoma and modulated PI3K/AKT signaling pathway. Arch Oral Biol 2019; 101: 85-91.
39. Torii T, Miyamoto Y, Nakamura K, Maeda M, Yamauchi J, Tanoue A. Arf6 guanine-nucleotide exchange factor, cytohesin-2, interacts with actinin-1 to regulate neurite extension. Cell Signal 2012; 24: 1872-1882.
40. LaMonica K, Ding HL, Artinger KB. prdm1a functions upstream of itga5 in zebrafish craniofacial development. Genesis 2015; 53: 270-277.
41. Chen J, Wang H, Wang J, Niu W, Deng C, Zhou M. LncRNA NEAT1 enhances glioma progression via regulating the miR-128-3p/ITGA5 axis. Mol Neurobiol 2021; 58: 5163-5177.
42. Lee YS, Yu JE, Kim MJ, Ham HJ, Jeon SH, Yun J, et al. New therapeutic strategy for atopic dermatitis by targeting CHI3L1/ITGA5 axis. Clin Transl Med 2022; 12: e739.
43. Xiong Y, Song X, Kudusi, Zu X, Chen M, He W, et al. Oncogenic GBX2 promotes the malignant behaviors of bladder cancer cells by binding to the ITGA5 promoter and activating its transcription. Funct Integr Genomics 2022; 22: 937-950.