Tweety homolog 3 promotes colorectal cancer progression through mutual regulation of histone deacetylase 7

Abstract

Colorectal cancer (CRC) is one of the leading cancers worldwide, with metastasis being a major cause of high mortality rates among patients. In this study, dysregulated gene Tweety homolog 3 (TTYH3) was identified by Gene Expression Omnibus database. Public databases were used to predict potential competing endogenous RNAs (ceRNAs) for TTYH3. Quantitative real-time polymerase chain reaction, western blot, and immunohistochemistry were utilized to analyze TTYH3 and histone deacetylase 7 (HDAC7) levels. Luciferase assays confirmed miR-1271-5p directly targeting the 3′ untranslated regions of TTYH3 and HDAC7. In vitro experiments such as transwell and human umbilical vein endothelial cell tube formation, as well as in vivo mouse models, were conducted to assess the biological functions of TTYH3 and HDAC7. We discovered that upregulation of TTYH3 in CRC promotes cell migration by affecting the Epithelial–mesenchymal transition pathway, which was independent of its ion channel activity. Mechanistically, TTYH3 and HDAC7 functioned as ceRNAs, reciprocally regulating each other's expression. TTYH3 competes for binding miR-1271-5p, increasing HDAC7 expression, facilitating CRC metastasis and angiogenesis. This study reveals the critical role of TTYH3 in promoting CRC metastasis through ceRNA crosstalk, offering new insights into potential therapeutic targets for clinical intervention.

Keywords

ceRNA / colorectal cancer / HDAC7 / migration / TTYH3

References

1	H Sung, J Ferlay, RL Siegel, et al. Global Cancer Statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2021;71(3):209-249.
2	RL Siegel, NS Wagle, A Cercek, RA Smith, A Jemal. Colorectal cancer statistics, 2023. CA Cancer J Clin. 2023;73(3):233-254.
3	LH Biller, D Schrag. Diagnosis and treatment of metastatic colorectal cancer: a review. JAMA. 2021;325(7):669-685.
4	AD Halleran, M Sehdev, BA Rabe, RW Huyck, CC Williams, MS Saha. Characterization of tweety gene (ttyh1-3) expression in Xenopus laevis during embryonic development. Gene Expr Patterns. 2015;17(1):38-44.
5	YE Han, J Kwon, J Won, et al. Tweety-homolog (Ttyh) family encodes the pore-forming subunits of the swelling-dependent volume-regulated anion channel (VRAC(swell)) in the brain. Exp Neurobiol. 2019;28(2):183-215.
6	Y He, AJ Ramsay, ML Hunt, AK Whitbread, SA Myers, JD Hooper. N-glycosylation analysis of the human Tweety family of putative chloride ion channels supports a penta-spanning membrane arrangement: impact of N-glycosylation on cellular processing of Tweety homologue 2 (TTYH2). Biochem J. 2008;412(1):45-55.
7	RR Nalamalapu, M Yue, AR Stone, S Murphy, MS Saha. The tweety gene family: from embryo to disease. Front Mol Neurosci. 2021;14:672511.
8	A Sukalskaia, MS Straub, D Deneka, M Sawicka, R Dutzler. Cryo-EM structures of the TTYH family reveal a novel architecture for lipid interactions. Nat Commun. 2021;12(1):4893.
9	M Suzuki. The Drosophila tweety family: molecular candidates for large-conductance Ca2+-activated Cl- channels. Exp Physiol. 2006;91(1):141-147.
10	M Suzuki, A Mizuno. A novel human Cl(-) channel family related to Drosophila flightless locus. J Biol Chem. 2004;279(21):22461-22468.
11	FK Rae, JD Hooper, HJ Eyre, GR Sutherland, DL Nicol, JA Clements. TTYH2, a human homologue of the Drosophila melanogaster gene tweety, is located on 17q24 and upregulated in renal cell carcinoma. Genomics. 2001;77(3):200-207.
12	Y Toiyama, A Mizoguchi, K Kimura, et al. TTYH2, a human homologue of the Drosophila melanogaster gene tweety, is up-regulated in colon carcinoma and involved in cell proliferation and cell aggregation. World J Gastroenterol. 2007;13(19):2717-2721.
13	DK Moon, YJ Bae, GR Jeong, CH Cho, SC Hwang. Upregulated TTYH2 expression is critical for the invasion and migration of U2OS human osteosarcoma cell lines. Biochem Biophys Res Commun. 2019;516(2):521-525.
14	Y Wang, Y Xie, B Dong, et al. The TTYH3/MK5 positive feedback loop regulates tumor progression via GSK3-β/β-catenin signaling in HCC. Int J Biol Sci. 2022;18(10):4053-4070.
15	PK Biswas, Y Kwak, A Kim, et al. TTYH3 modulates bladder cancer proliferation and metastasis via FGFR1/H-Ras/A-Raf/MEK/ERK pathway. Int J Mol Sci. 2022;23(18):10496.
16	FA Karreth, PP Pandolfi. ceRNA cross-talk in cancer: when ce-bling rivalries go awry. Cancer Discov. 2013;3(10):1113-1121.
17	Y Wang, J Hou, D He, et al. The emerging function and mechanism of ceRNAs in cancer. Trends Genet. 2016;32(4):211-224.
18	X Qi, Y Lin, J Chen, B Shen. Decoding competing endogenous RNA networks for cancer biomarker discovery. Brief Bioinform. 2020;21(2):441-457.
19	L Salmena, L Poliseno, Y Tay, L Kats, PP Pandolfi. A ceRNA hypothesis: the Rosetta Stone of a hidden RNA language? Cell. 2011;146(3):353-358.
20	Y Tay, L Kats, L Salmena, et al. Coding-independent regulation of the tumor suppressor PTEN by competing endogenous mRNAs. Cell. 2011;147(2):344-357.
21	N Prevarskaya, R Skryma, Y Shuba. Ion channels in cancer: are cancer hallmarks oncochannelopathies? Physiol Rev. 2018;98(2):559-621.
22	S Gao, H Liu, S Hou, et al. MiR-489 suppresses tumor growth and invasion by targeting HDAC7 in colorectal cancer. Clin Transl Oncol. 2018;20(6):703-712.
23	TP Chendrimada, RI Gregory, E Kumaraswamy, et al. TRBP recruits the Dicer complex to Ago2 for microRNA processing and gene silencing. Nature. 2005;436(7051):740-744.
24	L Wang, M Xu, CY Kao, SY Tsai, MJ Tsai. Small molecule JQ1 promotes prostate cancer invasion via BET-independent inactivation of FOXA1. J Clin Invest. 2020;130(4):1782-1792.
25	P Peixoto, A Blomme, B Costanza, et al. HDAC7 inhibition resets STAT3 tumorigenic activity in human glioblastoma independently of EGFR and PTEN: new opportunities for selected targeted therapies. Oncogene. 2016;35(34):4481-4494.
26	M To, S Yamamura, K Akashi, et al. Defect of adaptation to hypoxia in patients with COPD due to reduction of histone deacetylase 7. Chest. 2012;141(5):1233-1242.
27	W Xue, B Dong, Y Zhao, et al. Upregulation of TTYH3 promotes epithelial-to-mesenchymal transition through Wnt/β-catenin signaling and inhibits apoptosis in cholangiocarcinoma. Cell Oncol (Dordr). 2021;44(6):1351-1361.
28	U Banderali, L Leanza, N Eskandari, S Gentile. Potassium and chloride ion channels in cancer: a novel paradigm for cancer therapeutics. Rev Physiol Biochem Pharmacol. 2022;183:135-155.
29	K Guo, K Qian, Y Shi, T Sun, Z Wang. LncRNA-MIAT promotes thyroid cancer progression and function as ceRNA to target EZH2 by sponging miR-150-5p. Cell Death Dis. 2021;12(12):1097.
30	M Dokmanovic, G Perez, W Xu, et al. Histone deacetylase inhibitors selectively suppress expression of HDAC7. Mol Cancer Ther. 2007;6(9):2525-2534.
31	C Caslini, S Hong, YJ Ban, XS Chen, TA Ince. HDAC7 regulates histone 3 lysine 27 acetylation and transcriptional activity at super-enhancer-associated genes in breast cancer stem cells. Oncogene. 2019;38(39):6599-6614.
32	QG Li, T Xiao, W Zhu, et al. HDAC7 promotes the oncogenicity of nasopharyngeal carcinoma cells by miR-4465-EphA2 signaling axis. Cell Death Dis. 2020;11(5):322.
33	D Hanahan. Hallmarks of cancer: new dimensions. Cancer Discov. 2022;12(1):31-46.
34	Y Feng, Z Ma, M Pan, et al. WNT5A promotes the metastasis of esophageal squamous cell carcinoma by activating the HDAC7/SNAIL signaling pathway. Cell Death Dis. 2022;13(5):480.
35	S Chang, BD Young, S Li, X Qi, JA Richardson, EN Olson. Histone deacetylase 7 maintains vascular integrity by repressing matrix metalloproteinase 10. Cell. 2006;126(2):321-334.
36	D Mottet, A Bellahcène, S Pirotte, et al. Histone deacetylase 7 silencing alters endothelial cell migration, a key step in angiogenesis. Circ Res. 2007;101(12):1237-1246.
37	Y Pan, J Yang, Y Wei, et al. Histone deacetylase 7-derived peptides play a vital role in vascular repair and regeneration. Adv Sci (Weinh). 2018;5(8):1800006.
38	A Turtoi, D Mottet, N Matheus, et al. The angiogenesis suppressor gene AKAP12 is under the epigenetic control of HDAC7 in endothelial cells. Angiogenesis. 2012;15(4):543-554.
39	H Kato, S Tamamizu-Kato, F Shibasaki. Histone deacetylase 7 associates with hypoxia-inducible factor 1alpha and increases transcriptional activity. J Biol Chem. 2004;279(40):41966-41974.
40	B Zhang, J Chen, M Cui, Y Jiang. LncRNA ZFAS1/miR-1271-5p/HK2 promotes glioma development through regulating proliferation, migration, invasion and apoptosis. Neurochem Res. 2020;45(12):2828-2839.
41	Y Wang, Z Wang, K Cheng, Q Hao. FAM201A promotes cervical cancer progression and metastasis through miR-1271-5p/flotillin-1 axis targeting-induced Wnt/β-catenin pathway. J Oncol. 2022;2022:1123839.
42	T Li, Y Li, Y Gan, et al. Methylation-mediated repression of MiR-424/503 cluster promotes proliferation and migration of ovarian cancer cells through targeting the hub gene KIF23. Cell Cycle. 2019;18(14):1601-1618.
43	Y Li, J Liu, Q Xiao, et al. EN2 as an oncogene promotes tumor progression via regulating CCL20 in colorectal cancer. Cell Death Dis. 2020;11(7):604.
44	N Weidner, J Folkman, F Pozza, et al. Tumor angiogenesis: a new significant and independent prognostic indicator in early-stage breast carcinoma. J Natl Cancer Inst. 1992;84(24):1875-1887.