Volume 7, Issue 3 (2022)
SJMR 2022, 7(3): 177-193 |
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Rojhannezhad M, mowla S J, mohammad soltani B, vasei M, Taghizadeh S, Mirsadeghi L. Functional and Bioinformatic Investigation of the Regulatory Region of HER2-Associated Enhancer GH17J039694, HER2-Enhancer1, in Breast Cancer (SKBR3 and MCF7) and non-Breast Cancer Cell Lines (HEK293) Using CRISPR/Cas9 Technology. SJMR 2022; 7 (3) : 7
URL: http://saremjrm.com/article-1-277-en.html
URL: http://saremjrm.com/article-1-277-en.html
Mahdieh Rojhannezhad1 
, Seyed javad Mowla * 2, Bahram Mohammad soltani3 , Mohammad Vasei4 , Sara Taghizadeh5 , Leila Mirsadeghi6
, Seyed javad Mowla * 2, Bahram Mohammad soltani3 , Mohammad Vasei4 , Sara Taghizadeh5 , Leila Mirsadeghi6
1- PhD student in molecular genetics, Tarbiat Modares University, Tehran, Iran
2- Professor of Genetics, Department of Molecular Genetics, Tarbiat Modares University, Tehran, Iran ,sjmowla@modares.ac.ir
3- Professor of Genetics, Department of Molecular Genetics, Tarbiat Modares University, Tehran, Iran
4- Department of Pathology, Shariati Hospital, Tehran University of Medical Sciences, Iran
5- Pathology Laboratory, Shariati Hospital, Tehran University of Medical Sciences, Iran
6- Department of Bioinformatics, Institute of Biochemistry and Biophysics, university of Tehran, Iran
2- Professor of Genetics, Department of Molecular Genetics, Tarbiat Modares University, Tehran, Iran ,
3- Professor of Genetics, Department of Molecular Genetics, Tarbiat Modares University, Tehran, Iran
4- Department of Pathology, Shariati Hospital, Tehran University of Medical Sciences, Iran
5- Pathology Laboratory, Shariati Hospital, Tehran University of Medical Sciences, Iran
6- Department of Bioinformatics, Institute of Biochemistry and Biophysics, university of Tehran, Iran
Abstract: (1438 Views)
troduction: Her2 / neu gene (HER2) is a member of the family of epidermal growth factor receptors (EGFR) encoding a 185 kd protein with tyrosine kinase activity. This gene is expressed in a variety of tissues such as kidney, skin, colon, ovary and mammary glands and mostly is found in plasma membrane. Following amplifying or increasing of HER2 transcription, carcinogenesis or deterioration of tumorigenicity has been observed in some cancers. Epigenetic mechanisms and chromatin modifications play a significant role in gene expression, transcriptional regulation, and cancer progression. Studies have shown that these changes can also affect the sequence of enhancers, short DNA motifs that serve as binding sites for specific transcription factors. Chromatin regions such as DNase I sensitive sites and histone modifications such as H3K4me1 and H3K27ac are used to predict possible enhancers. Active enhancers that express eRNAs (non-coding transcriptional RNAs derived from enhancers) have two H3K4me1 and H3K4me3 histone marks. A therapeutic approach that directly targets genomic changes can be valuable because it has no effect on healthy cells. The development of genome editing tools such as the CRISPR system could provide such an opportunity. In this study, we aimed to investigate the regulatory role of the GH17J039694 region as an enhancer, which we called HER2-En1 (HER2-Enhancer1) for short, located in the HER2 gene in the 17q12 region: 37850592-37853472 (GRCh37/hg19.2009) with the help of bioinformatics studies and by using CRISPR/Cas9 technology for genetic manipulation.
Materials and Methods: Genetic editing of this region was done for cis and trans studies in HER2+ and HER2- breast cancer cells. Bioinformatics studies were conducted in different cell lines with the help of databases such as GEO gene, ChIP-Atlas and TCGA. The laboratory results showed the reduction of HER2 variants and expression changes of other studied genes. In general, according to bioinformatics and laboratory studies, it seems that this part of the HER2 gene can be considered as an integral regulatory region.
Results: The results showed the decrease in the expression of the variants in the studied cell lines after 24 hours in cis and the expression changes of other studied genes in trans. Also, bioinformatic studies such as histone modifications (H3K27ac) showed the importance of HER2-En1 as a regulatory region.
Conclusion: In general, according to bioinformatics and laboratory studies in the form of gene editing of the HER2-En1 region, it seems that this part of the HER2 sequence can be considered as an enhancer regulatory region.
Materials and Methods: Genetic editing of this region was done for cis and trans studies in HER2+ and HER2- breast cancer cells. Bioinformatics studies were conducted in different cell lines with the help of databases such as GEO gene, ChIP-Atlas and TCGA. The laboratory results showed the reduction of HER2 variants and expression changes of other studied genes. In general, according to bioinformatics and laboratory studies, it seems that this part of the HER2 gene can be considered as an integral regulatory region.
Results: The results showed the decrease in the expression of the variants in the studied cell lines after 24 hours in cis and the expression changes of other studied genes in trans. Also, bioinformatic studies such as histone modifications (H3K27ac) showed the importance of HER2-En1 as a regulatory region.
Conclusion: In general, according to bioinformatics and laboratory studies in the form of gene editing of the HER2-En1 region, it seems that this part of the HER2 sequence can be considered as an enhancer regulatory region.
Article number: 7
Article Type: Original Research |
Subject:
Women Diseases
Received: 2022/11/16 | Accepted: 2022/12/21 | Published: 2023/07/8
Received: 2022/11/16 | Accepted: 2022/12/21 | Published: 2023/07/8
References
1. Mungamuri, S.K., et al., Chromatin modifications sequentially enhance ErbB2 expression in ErbB2-positive breast cancers. Cell reports, 2013. 5(2): p. 302-313. [DOI:10.1016/j.celrep.2013.09.009]
2. Moasser, M.M., The oncogene HER2: its signaling and transforming functions and its role in human cancer pathogenesis. Oncogene, 2007. 26(45): p. 6469-6487. [DOI:10.1038/sj.onc.1210477]
3. Sirkisoon, S.R., et al., EGFR and HER2 signaling in breast cancer brain metastasis. Frontiers in bioscience (Elite edition), 2016. 8: p. 245. [DOI:10.2741/e765]
4. Choudhury, A., et al., Small interfering RNA (siRNA) inhibits the expression of the Her2/neu gene, upregulates HLA class I and induces apoptosis of Her2/neu positive tumor cell lines. International journal of cancer, 2004. 108(1): p. 71-77. [DOI:10.1002/ijc.11497]
5. Korkaya, H. and M.S. Wicha, HER2 and breast cancer stem cells: more than meets the eye. Cancer research, 2013. 73(12): p. 3489-3493. [DOI:10.1158/0008-5472.CAN-13-0260]
6. El Hadi, H., et al., Development and evaluation of a novel RT-qPCR based test for the quantification of HER2 gene expression in breast cancer. Gene, 2017. 605: p. 114-122. [DOI:10.1016/j.gene.2016.12.027]
7. Thariat, J. and P.-Y. Marcy, Neck dissection and chemoradiation in head and neck cancer. The Lancet Oncology, 2010. 11(3): p. 224-225. [DOI:10.1016/S1470-2045(10)70002-4]
8. Ménard, S., et al., Role of HER2 gene overexpression in breast carcinoma. Journal of cellular physiology, 2000. 182(2): p. 150-162.
https://doi.org/10.1002/(SICI)1097-4652(200002)182:2<150::AID-JCP3>3.0.CO;2-E [DOI:10.1002/(SICI)1097-4652(200002)182:23.0.CO;2-E]
9. Merry, C.R., et al., Transcriptome-wide identification of mRNAs and lincRNAs associated with trastuzumab-resistance in HER2-positive breast cancer. Oncotarget, 2016. 7(33): p. 53230. [DOI:10.18632/oncotarget.10637]
10. Shlyueva, D., G. Stampfel, and A. Stark, Transcriptional enhancers: from properties to genome-wide predictions. Nature Reviews Genetics, 2014. 15(4): p. 272-286. [DOI:10.1038/nrg3682]
11. Zlotorynski, E., Gene expression: Developmental enhancers in action. Nature Reviews Genetics, 2018. 19(4): p. 187. [DOI:10.1038/nrg.2018.13]
12. Li, W., D. Notani, and M.G. Rosenfeld, Enhancers as non-coding RNA transcription units: recent insights and future perspectives. Nature Reviews Genetics, 2016. 17(4): p. 207-223. [DOI:10.1038/nrg.2016.4]
13. Shin, H.Y., Targeting super-enhancers for disease treatment and diagnosis. Molecules and cells, 2018. 41(6): p. 506.
14. Nebbioso, A., et al., Cancer epigenetics: moving forward. PLoS genetics, 2018. 14(6): p. e1007362. [DOI:10.1371/journal.pgen.1007362]
15. Xi, Y., et al., Histone modification profiling in breast cancer cell lines highlights commonalities and differences among subtypes. BMC genomics, 2018. 19(1): p. 1-11. [DOI:10.1186/s12864-018-4533-0]
16. Liu, Q., et al., A novel HER2 gene body enhancer contributes to HER2 expression. Oncogene, 2018. 37(5): p. 687-694. [DOI:10.1038/onc.2017.382]
17. Wang, H. and W. Sun, CRISPR-mediated targeting of HER2 inhibits cell proliferation through a dominant negative mutation. Cancer letters, 2017. 385: p. 137-143. [DOI:10.1016/j.canlet.2016.10.033]
18. Huang, D.W., B.T. Sherman, and R.A. Lempicki, Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nature protocols, 2009. 4(1): p. 44-57. [DOI:10.1038/nprot.2008.211]
19. Viewer, I.G., Broad Institute. URL: https://software. broadinstitute. org/software/igv/[accessed 2022-05-12].
20. Wilks, C., et al., recount3: summaries and queries for large-scale RNA-seq expression and splicing. Genome biology, 2021. 22(1): p. 1-40. [DOI:10.1186/s13059-021-02533-6]
21. Oki, S., et al., Ch IP‐Atlas: a data‐mining suite powered by full integration of public Ch IP‐seq data. EMBO reports, 2018. 19(12): p. e46255. [DOI:10.15252/embr.201846255]
22. Chao, W.-R., et al., HER2 amplification and overexpression are significantly correlated in mucinous epithelial ovarian cancer. Human pathology, 2014. 45(4): p. 810-816. [DOI:10.1016/j.humpath.2013.11.016]
23. Nencioni, A., et al., Grb7 upregulation is a molecular adaptation to HER2 signaling inhibition due to removal of Akt-mediated gene repression. PloS one, 2010. 5(2): p. e9024. [DOI:10.1371/journal.pone.0009024]
24. Manning, B.D. and L.C. Cantley, AKT/PKB signaling: navigating downstream. Cell, 2007. 129(7): p. 1261-1274. [DOI:10.1016/j.cell.2007.06.009]
25. Hemmings, B.A. and D.F. Restuccia, Pi3k-pkb/akt pathway. Cold Spring Harbor perspectives in biology, 2012. 4(9): p. a011189. [DOI:10.1101/cshperspect.a011189]
26. Szklarczyk, D., et al., The STRING database in 2017: quality-controlled protein-protein association networks, made broadly accessible. Nucleic acids research, 2016: p. gkw937. [DOI:10.1093/nar/gkw937]
27. Stanimir, M., et al., Mullerianosis of the urinary bladder: a rare case report and review of the literature. Rom J Morphol Embryol, 2016. 57(2 Suppl): p. 849-852.
28. Klann, T.S., et al., CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome. Nature biotechnology, 2017. 35(6): p. 561-568. [DOI:10.1038/nbt.3853]
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