22Aug 2017

CRISPR KNOCKOUT OF TAX1BP3 REDUCES EXPRESSION OF P53 IN HUMAN PAPILLOMA NEGATIVE AND HUMAN PAPILLOMA POSITIVE CERVICAL CARCINOMA CELLS.

  • Department of Medical Biochemistry, Faculty of Medicine, Mansoura University, El Gomhouria Street, Dakahlia Governorate, Egypt.
  • Department of Obstetrics and Gynecology, Faculty of Medicine, Mansoura University, El Gomhouria Street, Dakahlia Governorate, Egypt.
  • School of Cancer and Imaging Science, Research Floor, St Mary's Hospital, Hathersage Road, Manchester M13 0JH. UK.
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Background: The PDZ protein Tax 1 binding protein 3 (Tax1BP3) is unique since, unlike other PDZ proteins, it contains a single rather than multiple PDZ domains. Previous work has shown Tax1BP3 is a target of oncoproteins from two separate human tumor viruses and it is known to regulate Rho and wnt/β-catenin signaling. Most importantly Tax1BP3 has been shown to promote degradation of the p53 protein in glioma cells although it is not clear whether it performs this same function in tumor virus positive cell types . Methods: The cervical cancer cell lines C33A (HPV negative), C33AE6 (HPV16 E6 transfected) and CaSki (HPV16 positive) were stably transfected with a CRISPR/Cas9 Tax1BP3 knockout construct. Levels of Tax1BP3 mRNA were initially determined in puromycin resistant monoclonal cells by RT-PCR. Tax1BP3 and p53 protein levels were then analyzed by a combination of western immunoblotting and flow cytometry. Results: Both bi and monoallelic Tax1BP3 targeted cell lines were isolated for all three cell types which showed either partial or complete silencing of Tax1BP3 expression. Irrespective of cell type, all cells which had reduced or absent Tax1BP3 expression showed much lower levels of p53 protein than control cells.. Conclusion: Unlike previous findings with glioma cells, the presented data showed that reduced levels of Tax1BP3 directly correlated with reduced levels of p53 protein expression. Furthermore, this effect was seen in cells whether or not they expressed mutant (C33A) or wild type (CaSki) p53 or full or partial HPV genome (CaSki and C33AE6). These results suggest that Tax1BP3 regulates p53 protein levels in HPV positive and negative cells by a novel and non-canonical means.

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[Mona El Helaly El Sherbeny, Hussein Abdelaziz Abdalla, El Said Mohamed Abd El Hady, Ian Noel Hampson, Anthony.W.Oliver, Souad Mohamed Abo Azma and Muhamad Muhamad Said. (2017); CRISPR KNOCKOUT OF TAX1BP3 REDUCES EXPRESSION OF P53 IN HUMAN PAPILLOMA NEGATIVE AND HUMAN PAPILLOMA POSITIVE CERVICAL CARCINOMA CELLS. Int. J. of Adv. Res. 5 (Aug). 1402-1411] (ISSN 2320-5407). www.journalijar.com

Mona El Helaly El Sherbeny
Mansoura university, Faculty of Medicine, Medical biochemistry and molecular biology department, El Gomhouria Street, Dakahlia Governorate , Egypt

DOI:

Article DOI: 10.21474/IJAR01/5197      
DOI URL: https://dx.doi.org/10.21474/IJAR01/5197

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