HBXIP knockdown inhibits FHL2 to promote cycle arrest and suppress cervical cancer cell proliferation, invasion and migration

Gao,Xia; Yang,Lina

doi:10.3892/ol.2023.13772

HBXIP knockdown inhibits FHL2 to promote cycle arrest and suppress cervical cancer cell proliferation, invasion and migration

Authors:
- Xia Gao
- Lina Yang
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Affiliations: Department of Gynaecology, Heping Hospital Affiliated to Changzhi Medical College, Changzhi, Shanxi 046000, P.R. China, Department of Gynecology, The 521 Hospital of Norinco Group, Xi'an, Shaanxi 710065, P.R. China
Published online on: March 24, 2023 https://doi.org/10.3892/ol.2023.13772
Article Number: 186
Copyright: © Gao et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Hepatitis B X‑interacting protein (HBXIP) and four and a half LIM domain 2 (FHL2) have been reported to serve as independent biomarkers for cervical cancer. The present study evaluated the effects of HBXIP on cervical cancer in terms of its cellular malignant characteristics. Reverse transcription‑quantitative PCR and western blotting were used to assess the mRNA and protein expression levels of HBXIP and FHL2 in the human endocervical epithelial End1/E6E7 cell line and the cervical cancer HeLa, CaSki, C33A and SiHa cell lines. After knocking down HBXIP expression by transfection of small interfering RNAs targeting HBXIP, cell cycle progression was assessed using flow cytometry with PI staining. Cell Counting Kit‑8, 5‑ethynyl‑2'‑deoxyuridine staining, wound healing and Transwell assays were used to assess cell proliferation, migration and invasion, respectively. Furthermore, co‑immunoprecipitation assay was used to evaluate the potential binding relationship between HBXIP and FHL2. Western blotting was used for the analysis of HBXIP and FHL2, cell cycle‑associated proteins, including cyclin D1 and cyclin D2, metastasis‑associated proteins, including MMP2 and MMP9, and Wnt/β‑catenin signaling‑associated proteins, including β‑catenin and c‑Myc. Both HBXIP and FHL2 were found to be highly expressed in cervical cancer cells compared with that in the human endocervical epithelial cell line. HBXIP knockdown suppressed the proliferation, invasion and migration of HeLa cells, but promoted cell cycle arrest at the G0/G1 phase. HBXIP was demonstrated to interact with FHL2, and HBXIP knockdown also inhibited FHL2 mRNA and protein expression. By contrast, FHL2 overexpression reversed the inhibitory effects of HBXIP knockdown on the malignant characteristics of cervical cancer cells. Furthermore, HBXIP knockdown blocked the Wnt/β‑catenin signaling pathway in HeLa cells, which was also partially reversed by FHL2 overexpression; the decreased β‑catenin and c‑Myc expression caused by HBXIP knockdown was increased again after FHL2 was overexpressed. In conclusion, these results suggest that HBXIP knockdown suppressed the malignant characteristics of cervical cancer cells through the downregulation of FHL2 expression, indicating a promising insight into the therapeutic target of cervical cancer.

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1	Lopez MS, Baker ES, Maza M, Fontes-Cintra G, Lopez A, Carvajal JM, Nozar F, Fiol V and Schmeler KM: Cervical cancer prevention and treatment in Latin America. J Surg Oncol. 115:615–618. 2017. View Article : Google Scholar : PubMed/NCBI
2	Canfell K, Kim JJ, Brisson M, Keane A, Simms KT, Caruana M, Burger EA, Martin D, Nguyen DTN, Bénard É, et al: Mortality impact of achieving WHO cervical cancer elimination targets: A comparative modelling analysis in 78 low-income and lower-middle-income countries. Lancet. 395:591–603. 2020. View Article : Google Scholar : PubMed/NCBI
3	Johnson CA, James D, Marzan A and Armaos M: Cervical cancer: An overview of pathophysiology and management. Semin Oncol Nurs. 35:166–174. 2019. View Article : Google Scholar : PubMed/NCBI
4	Zimet GD, Rosberger Z, Fisher WA, Perez S and Stupiansky NW: Beliefs, behaviors and HPV vaccine: Correcting the myths and the misinformation. Prev Med. 57:414–418. 2013. View Article : Google Scholar : PubMed/NCBI
5	Yang Z and Jiang X, Li D and Jiang X: HBXIP promotes gastric cancer via METTL3-mediated MYC mRNA m6A modification. Aging (Albany NY). 12:24967–24982. 2020. View Article : Google Scholar : PubMed/NCBI
6	Zheng S, Wu H, Wang F, Lv J, Lu J, Fang Q, Wang F, Lu Y, Zhang S, Xu Y, et al: The oncoprotein HBXIP facilitates metastasis of hepatocellular carcinoma cells by activation of MMP15 expression. Cancer Manag Res. 11:4529–4540. 2019. View Article : Google Scholar : PubMed/NCBI
7	Zhao Y, Li H, Zhang Y, Li L, Fang R, Li Y, Liu Q, Zhang W, Qiu L, Liu F, et al: Oncoprotein HBXIP modulates abnormal lipid metabolism and growth of breast cancer cells by activating the LXRs/SREBP-1c/FAS signaling cascade. Cancer Res. 76:4696–4707. 2016. View Article : Google Scholar : PubMed/NCBI
8	Qiu L, Lu F, Zhang L, Wang G, Geng R and Miao Y: HBXIP regulates gastric cancer glucose metabolism and malignancy through PI3K/AKT and p53 signaling. Onco Targets Ther. 13:3359–3374. 2020. View Article : Google Scholar : PubMed/NCBI
9	Wang X, Feng Q, Yu H, Zhou X, Shan C, Zhang Q and Liu S: HBXIP: A potential prognosis biomarker of colorectal cancer which promotes invasion and migration via epithelial-mesenchymal transition. Life Sci. 245:1173542020. View Article : Google Scholar : PubMed/NCBI
10	Xiu M, Zeng X, Shan R, Wen W, Li J and Wan R: The oncogenic role of HBXIP. Biomed Pharmacother. 133:1110452021. View Article : Google Scholar : PubMed/NCBI
11	Li N, Wang Y, Che S, Yang Y, Piao J, Liu S and Lin Z: HBXIP over expression as an independent biomarker for cervical cancer. Exp Mol Pathol. 102:133–137. 2017. View Article : Google Scholar : PubMed/NCBI
12	Chan KK, Tsui SK, Lee SM, Luk SC, Liew CC, Fung KP, Waye MM and Lee CY: Molecular cloning and characterization of FHL2, a novel LIM domain protein preferentially expressed in human heart. Gene. 210:345–350. 1998. View Article : Google Scholar : PubMed/NCBI
13	Kinoshita M, Nakagawa T, Shimizu A and Katsuoka Y: Differently regulated androgen receptor transcriptional complex in prostate cancer compared with normal prostate. Int J Urol. 12:390–397. 2005. View Article : Google Scholar : PubMed/NCBI
14	Genini M, Schwalbe P, Scholl FA, Remppis A, Mattei MG and Schäfer BW: Subtractive cloning and characterization of DRAL, a novel LIM-domain protein down-regulated in rhabdomyosarcoma. DNA Cell Biol. 16:433–442. 1997. View Article : Google Scholar : PubMed/NCBI
15	Xu J, Zhou J, Li MS, Ng CF, Ng YK, Lai PBS and Tsui SKW: Transcriptional regulation of the tumor suppressor FHL2 by p53 in human kidney and liver cells. PLoS One. 9:e993592014. View Article : Google Scholar : PubMed/NCBI
16	Gabriel B, Mildenberger S, Weisser CW, Metzger E, Gitsch G, Schüle R and Müller JM: Focal adhesion kinase interacts with the transcriptional coactivator FHL2 and both are overexpressed in epithelial ovarian cancer. Anticancer Res. 24:921–927. 2004.PubMed/NCBI
17	Chan KK, Tsui SK, Ngai SM, Lee SM, Kotaka M, Waye MM, Lee CY and Fung KP: Protein-protein interaction of FHL2, a LIM domain protein preferentially expressed in human heart, with hCDC47. J Cell Biochem. 76:499–508. 2000. View Article : Google Scholar : PubMed/NCBI
18	Chen D, Xu W, Bales E, Colmenares C, Conacci-Sorrell M, Ishii S, Stavnezer E, Campisi J, Fisher DE, Ben-Ze'ev A and Medrano EE: SKI activates Wnt/beta-catenin signaling in human melanoma. Cancer Res. 63:6626–6634. 2003.PubMed/NCBI
19	Jin H, Lee K, Kim YH, Oh HK, Maeng YI, Kim TM, Suh DS and Bae J: Scaffold protein FHL2 facilitates MDM2-mediated degradation of IER3 to regulate proliferation of cervical cancer cells. Oncogene. 35:5106–5118. 2016. View Article : Google Scholar : PubMed/NCBI
20	Jin X, Jiao X, Jiao J, Zhang T and Cui B: Increased expression of FHL2 promotes tumorigenesis in cervical cancer and is correlated with poor prognosis. Gene. 669:99–106. 2018. View Article : Google Scholar : PubMed/NCBI
21	Cao CY, Mok SWF, Cheng VWS and Tsui SKW: The FHL2 regulation in the transcriptional circuitry of human cancers. Gene. 572:1–7. 2015. View Article : Google Scholar : PubMed/NCBI
22	Brun J, Dieudonné FX, Marty C, Müller J, Schüle R, Patiño-García A, Lecanda F, Fromigué O and Marie PJ: FHL2 silencing reduces Wnt signaling and osteosarcoma tumorigenesis in vitro and in vivo. PLoS One. 8:e550342013. View Article : Google Scholar : PubMed/NCBI
23	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
24	Qie S and Diehl JA: Cyclin D1, cancer progression, and opportunities in cancer treatment. J Mol Med (Berl). 94:1313–1326. 2016. View Article : Google Scholar : PubMed/NCBI
25	Hamidi H and Ivaska J: Every step of the way: Integrins in cancer progression and metastasis. Nat Rev Cancer. 18:533–548. 2018. View Article : Google Scholar : PubMed/NCBI
26	Oudin MJ and Weaver VM: Physical and chemical gradients in the tumor microenvironment regulate tumor cell invasion, migration, and metastasis. Cold Spring Harb Symp Quant Biol. 81:189–205. 2016. View Article : Google Scholar : PubMed/NCBI
27	Nie H, Bu F, Xu J, Li T and Huang J: 29 immune-related genes pairs signature predict the prognosis of cervical cancer patients. Sci Rep. 10:141522020. View Article : Google Scholar : PubMed/NCBI
28	Wang FZ, Fei HR, Lian LH, Wang JM and Qiu YY: Hepatitis B x-interacting protein induces HepG2 cell proliferation through activation of the phosphatidylinositol 3-kinase/Akt pathway. Exp Biol Med (Maywood). 236:62–69. 2011. View Article : Google Scholar : PubMed/NCBI
29	Li Y, Zhang Z, Zhou X, Li L, Liu Q, Wang Z, Bai X, Zhao Y, Shi H, Zhang X and Ye L: The oncoprotein HBXIP enhances migration of breast cancer cells through increasing filopodia formation involving MEKK2/ERK1/2/Capn4 signaling. Cancer Lett. 355:288–296. 2014. View Article : Google Scholar : PubMed/NCBI
30	Wang Y, Cui M, Cai X, Sun B, Liu F, Zhang X and Ye L: The oncoprotein HBXIP up-regulates SCG3 through modulating E2F1 and miR-509-3p in hepatoma cells. Cancer Lett. 352:169–178. 2014. View Article : Google Scholar : PubMed/NCBI
31	Zheng K, He Z, Kitazato K and Wang Y: Selective autophagy regulates cell cycle in cancer therapy. Theranostics. 9:104–125. 2019. View Article : Google Scholar : PubMed/NCBI
32	Fei H, Zhou Y, Li R, Yang M, Ma J and Wang F: HBXIP, a binding protein of HBx, regulates maintenance of the G2/M phase checkpoint induced by DNA damage and enhances sensitivity to doxorubicin-induced cytotoxicity. Cell Cycle. 16:468–476. 2017. View Article : Google Scholar : PubMed/NCBI
33	Wang C, Lv X, He C, Davis JS, Wang C and Hua G: Four and a half LIM domains 2 (FHL2) contribute to the epithelial ovarian cancer carcinogenesis. Int J Mol Sci. 21:77512020. View Article : Google Scholar : PubMed/NCBI
34	Hua G, He C, Lv X, Fan L, Wang C, Remmenga SW, Rodabaugh KJ, Yang L, Lele SM, Yang P, et al: The four and a half LIM domains 2 (FHL2) regulates ovarian granulosa cell tumor progression via controlling AKT1 transcription. Cell Death Dis. 7:e22972016. View Article : Google Scholar : PubMed/NCBI
35	Shao C, Qiu Y, Liu J, Feng H, Shen S, Saiyin H, Yu W, Wei Y, Yu L, Su W and Wu J: PARP12 (ARTD12) suppresses hepatocellular carcinoma metastasis through interacting with FHL2 and regulating its stability. Cell Death Dis. 9:8562018. View Article : Google Scholar : PubMed/NCBI
36	Duchartre Y, Kim YM and Kahn M: The Wnt signaling pathway in cancer. Crit Rev Oncol Hematol. 99:141–149. 2016. View Article : Google Scholar : PubMed/NCBI
37	Bahrami A, Amerizadeh F, ShahidSales S, Khazaei M, Ghayour-Mobarhan M, Sadeghnia HR, Maftouh M, Hassanian SM and Avan A: Therapeutic potential of targeting wnt/beta-catenin pathway in treatment of colorectal cancer: Rational and progress. J Cell Biochem. 118:1979–1983. 2017. View Article : Google Scholar : PubMed/NCBI
38	Cai T, Sun D, Duan Y, Qiu Y, Dai C, Yang J and He W: FHL2 promotes tubular epithelial-to-mesenchymal transition through modulating beta-catenin signalling. J Cell Mol Med. 22:1684–1695. 2018. View Article : Google Scholar : PubMed/NCBI