--

12 (2) 2022

Establishing protocol for detecting methylation of Ubiquitin carboxyl terminal hydrolase 1 gene’s promoter in nasopharyngeal carcinoma


Author - Affiliation:
Thieu Hong Hue - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Huynh Thi Ngoc Tuyen - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Nguyen Van Truong - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Dao Duy Tin - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Tran Minh Dang Khoa - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Le Huyen Ai Thuy - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Lao Duc Thuan - Ho Chi Minh City Open University, Ho Chi Minh City , Vietnam
Corresponding author: Lao Duc Thuan - thuan.ld@ou.edu.vn
Submitted: 03-10-2022
Accepted: 24-10-2022
Published: 04-11-2022

Abstract
Background: The methylation of Ubiquitin Carboxyl Terminal Hydrolase 1 (UCHL1) gene’s promoter has been reported as the etiological factor of nasopharyngeal tumorigenesis. Purpose: This study is designed to establish a protocol for detecting methylation of UCHL1 gene’s promoter in nasopharyngeal carcinoma (NPC) in a Vietnamese population. Materials and methods: 10 samples of NPC biopsy tissues and 10 samples of non-cancerous swabs were collected from the local hospital. Chloroform/Phenol method and Nested-MSP assays were established to detect methylation of a target gene. Results: The isolated DNA reached purity and high concentration which were confirmed by the method of absorbance measurement at 260nm and 280nm. Additionally, the Nested-MSP products of methylation or unmethylation were analyzed and visualized in the agarose gel with the band of 169bps and 210bps, respectively. By sequencing, it was confirmed that the two sets of primer could distinguish the status of methylation and unmethylation of UCHL1 gene’s promoter. Conclusion: Our data suggested that the current protocol could successfully identify the status of methylation and/or unmethylation of UCHL1 gene’s promoter.

Keywords
epigenetics; methylation; Nested-MSP; Ubiquitin carboxyl terminal hydrolase 1; unmethylation

Full Text:
PDF

Cite this paper as:

Thieu, H. H., Huynh, T. T. N., Nguyen, T. V., Dao, T. D., Tran, K. M. D., Le, T. H. A., & Lao, T. D. (2022). Establishing protocol for detecting methylation of Ubiquitin carboxyl terminal hydrolase 1 gene’s promoter in nasopharyngeal carcinoma. Ho Chi Minh City Open University Journal of Science – Engineering and Technology, 12(2), 34-41. doi:10.46223/HCMCOUJS.tech.en.12.2.2542.2022


References

Dai, W., Zheng, H., Cheung, A. K., & Lung, M. L. (2016). Genetic and epigenetic landscape of nasopharyngeal carcinoma. Chinese Clinical Oncology, 5(2), Article 16. doi:10.21037/cco.2016.03.06


Divine, K. K., Liechty, K. C., Crume, K. C., & Belinsky, S. A. (2006). Nested multigene MSP/DHPLC method for analyzing promoter hypermethylation status in clinical samples. BioTechniques, 40(1), 40-48. doi:10.2144/000112095


Gong, B., Cao, Z., Zheng, P., Vitolo, O. V., Liu, S., Staniszewski, A., … Arancio, O. (2006). Ubiquitin hydrolase Uch-L1 rescues beta-amyloid-induced decreases in synaptic function and contextual memory. Cell, 126(4), 775-788. doi:10.1016/j.cell.2006.06.046


Goto, Y., Zeng, L., Yeom, C. J., Zhu, Y., Morinibu, A., Shinomiya, K., … Harada, H. (2015). UCHL1 provides diagnostic and antimetastatic strategies due to its deubiquitinating effect on HIF-1α. Nature Communications, 6(1), 1-13. doi:10.1038/ncomms7153


Han, B., Yang, X., Zhang, P., Zhang, Y., Tu, Y., He, Z., … Sun, H. (2020). DNA methylation biomarkers for nasopharyngeal carcinoma. PloS One, 15(4), Article e0230524. doi:10.1371/journal.pone.0230524


Herceg, Z., & Hainaut, P. (2007). Genetic and epigenetic alterations as biomarkers for cancer detection, diagnosis and prognosis. Molecular Oncology, 1(1), 26-41. doi:10.1016/j.molonc.2007.01.004


Hussain, S., Bedekovics, T., Liu, Q., Hu, W., Jeon, H., Johnson, S. H., … Galardy, P. J. (2018). UCH-L1 bypasses mTOR to promote protein biosynthesis and is required for MYC-driven lymphomagenesis in mice. Blood, The Journal of the American Society of Hematology, 132(24), 2564-2574. doi:10.1182/blood-2018-05-848515


Lao, T. D., Nguyen, T. N., & Le, T. (2021). Promoter hypermethylation of tumor suppressor genes located on short arm of the chromosome 3 as potential biomarker for the diagnosis of nasopharyngeal carcinoma. Diagnostics, 11(8), Article 1404. doi:10.3390/diagnostics11081404


Lao, T. D., Truong, P. K., Thieu, H. H., Nguyen, D. H., Nguyen, M. T., & Le, T. (2021). Simultaneously both expression of LMP-1 and methylation of e-cadherin: Molecular biomarker in stage IV of nasopharyngeal carcinoma patients. Balkan Journal of Medical Genetics: BJMG, 24(1), 57-66. doi:10.2478/bjmg-2021-0005


Li, L., Tao, Q., Jin, H., van Hasselt, A., Poon, F. F., Wang, X., … Cao, Y. (2010). The tumor suppressor UCHL1 forms a complex with p53/MDM2/ARF to promote p53 signaling and is frequently silenced in nasopharyngeal carcinoma. Clinical Cancer Research, 16(11), 2949-2958. doi:10.1158/1078-0432.CCR-09-3178


Licchesi, J. D., & Herman, J. G. (2009). Methylation-specific PCR. In DNA methylation (pp. 305-323). New York, NY: Humana Press.


Loyo, M., Brait, M., Kim, M. S., Ostrow, K. L., Jie, C. C., Chuang, A. Y., … Sidransky, D. (2011). A survey of methylated candidate tumor suppressor genes in nasopharyngeal carcinoma. International Journal of Cancer, 128(6), 1393-1403. doi:10.1002/ijc.25443


Moore, L. D., Le, T., & Fan, G. (2013). DNA methylation and its basic function. Neuropsychopharmacology, 38(1), 23-38. doi:10.1038/npp.2012.112


Nijman, S. M., Luna-Vargas, M. P., Velds, A., Brummelkamp, T. R., Dirac, A. M., Sixma, T. K., & Bernards, R. (2005). A genomic and functional inventory of deubiquitinating enzymes. Cell, 123(5), 773-786. doi:10.1016/j.cell.2005.11.007


Olson, N. D., & Morrow, J. B. (2012). DNA extract characterization process for microbial detection methods development and validation. BMC Research Notes, 5(1), 1-14. doi:10.1186/1756-0500-5-668


Sharma, S., Kelly, T. K., & Jones, P. A. (2010). Epigenetics in cancer. Carcinogenesis, 31(1), 27-36. doi:10.1093/carcin/bgp220


Tian, F., Yip, S. P., Kwong, D. L., Lin, Z., Yang, Z., & Wu, V. W. (2013). Promoter hypermethylation of tumor suppressor genes in serum as potential biomarker for the diagnosis of nasopharyngeal carcinoma. Cancer Epidemiology, 37(5), 708-713. doi:10.1016/j.canep.2013.05.012


Tsao, S. W., Yip, Y. L., Tsang, C. M., Pang, P. S., Lau, V. M., Zhang, G., & Lo, K. W. (2014). Etiological factors of nasopharyngeal carcinoma. Oral Oncology, 50(5), 330-338. doi:10.1016/j.oraloncology.2014.02.006


Wilfinger, W. W., Mackey, K., & Chomczynski, P. (2006). Assessing the quantity, purity and integrity of RNA and DNA following nucleic acid purification. In DNA sequencing II optimizing preparation and cleanup (vol. 291, p. 312). Burlington, MA: Jones and Bartlett Publishers.


Yu, J., Tao, Q., Cheung, K. F., Jin, H., Poon, F. F., Wang, X., … Sung, J. J. (2008). Epigenetic identification of ubiquitin carboxyl-terminal hydrolase L1 as a functional tumor suppressor and biomarker for hepatocellular carcinoma and other digestive tumors. Hepatology, 48(2), 508-518. doi:10.1002/hep.22343


Zhao, Y., Lei, Y., He, S. W., Li, Y. Q., Wang, Y. Q., Hong, X. H., … Tang, L. L. (2020). Hypermethylation of UCHL1 Promotes Metastasis of Nasopharyngeal Carcinoma by Suppressing Degradation of Cortactin (CTTN). Cells, 9(3), Article 559. doi:10.3390/cells9030559



DOI: https://doi.org/10.46223/HCMCOUJS.tech.en.12.2.2542.2022
Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.