Targeting HPV-16 antigens to the endoplasmic reticulum induces an endoplasmic reticulum stress response
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Very promising results have been observed with a deoxyribonucleic acid (DNA) vaccine based on human papillomavirus type-16 (HPV-16) antigen retention and delivery system in the endoplasmic reticulum (ER). However, the mechanism by which these antigens are processed once they reach this organelle is still unknown. Therefore, we evaluated whether this system awakens a stress response in the ER. Different DNA constructs based on E6 and E7 mutant antigens fused to an ER signal peptide (SP), a signal for retention in the ER (KDEL), or both signals (SPK), were transfected into HEK-293 cells. Overexpression of E6 and E7 antigens targeted to the ER (SP, and SPK constructs) induced ER stress, which was indicated by an increase of the ER-stress markers GRP78/BiP and CHOP. Additionally, the ER stress response was mediated by the ATF4 transcription factor, which was translocated into the nucleus. Besides, the overexpressed antigens were degraded by the proteasome. Through a cycloheximide-chase assay, we demonstrated that when both protein synthesis and proteasome were inhibited, the overexpressed antigens were degraded. Interestingly, when proteasome was blocked autophagy was increased and the ER stress response decreased. Taken together, these results indicate that the antigens are initially degraded by the ERAD pathway, and autophagy degradation pathway can be induced to compensate the proteasome inhibition. Therefore, we provided a new insight into the mechanism by which E6 and E7 mutant antigens are processed once they reach the ER, which will help to improve the development of more effective vaccines against cancer.
KeywordsE7 Stress response Signal peptide KDEL signal ER targeting GRP78/BiP
This study was supported through grants from the Programa de Apoyo a la Investigacion Cientifica y Tecnologica (grant no. SA179-15) from the Universidad Autonoma de Nuevo Leon and the National Council for Science and Technology (CONACYT; grant no. CB10-158509). DHMP was the recipient of a scholarship from CONACYT.
- Leach MR, Williams DB (2013) Calnexin and Calreticulin, Molecular Chaperones of the Endoplasmic Reticulum. Landes BioscienceGoogle Scholar
- Loera-Arias MJ, Martínez-Pérez AG, Barrera-Hernández A, Ibarra-Obregón ER, González-Saldívar G, Martínez-Ortega JI, Rosas-Taraco A, Villanueva-Olivo A, Esparza-González SC, Villatoro-Hernandez J, Saucedo-Cárdenas O, Montes-de-Oca-Luna R (2010) Targeting and retention of HPV16 E7 to the endoplasmic reticulum enhances immune tumour protection. J Cell Mol Med 14:890–894. https://doi.org/10.1111/j.1582-4934.2009.00934.x CrossRefGoogle Scholar
- Mak BC, Wang Q, Laschinger C, Lee W, Ron D, Harding HP, Kaufman RJ, Scheuner D, Austin RC, McCulloch CA (2008) Novel function of PERK as a mediator of force-induced apoptosis. J Biol Chem 283:23462–23472. https://doi.org/10.1074/jbc.M803194200
- Oslowski CM, Urano F (2011) Measuring ER stress and the unfolded protein response using mammalian tissue culture system. Methods Enzymol 490:71–92. https://doi.org/10.1016/B978-0-12-385114-7.00004-0 CrossRefGoogle Scholar
- Perez-Trujillo JJ, Garza-Morales R, Barron-Cantu JA, Figueroa-Parra G, Garcia-Garcia A, Rodriguez-Rocha H, Garcia-Juarez J, Muñoz-Maldonado GE, Saucedo-Cardenas O, Montes-de-Oca-Luna R, Loera-Arias MDJ (2017) DNA vaccine encoding human papillomavirus antigens flanked by a signal peptide and a KDEL sequence induces a potent therapeutic antitumor effect. Oncol Lett 13:1569–1574. https://doi.org/10.3892/ol.2017.5635 CrossRefGoogle Scholar
- Pérez-Trujillo JJ, Robles-Rodríguez OA, Garza-Morales R, García-García A, Rodríguez-Rocha H, Villanueva-Olivo A, Segoviano-Ramírez JC, Esparza-González SC, Saucedo-Cárdenas O, Montes-de-Oca-Luna R, Loera-Arias MJ (2018) Antitumor response by endoplasmic reticulum-targeting DNA vaccine is improved by adding a KDEL retention signal. Nucleic Acid Ther 28:252–261. https://doi.org/10.1089/nat.2017.0717
- Sherritt M, Cooper L, Moss DJ, Kienzle N, Altman J, Khanna R (2001) Immunization with tumor-associated epitopes fused to an endoplasmic reticulum translocation signal sequence affords protection against tumors with down-regulated expression of MHC and peptide transporters. Int Immunol 13:265–271CrossRefGoogle Scholar
- Shi W, Bu P, Liu J, Polack A, Fisher S, Qiao L (1999) Human papillomavirus type 16 E7 DNA vaccine: mutation in the open reading frame of E7 enhances specific cytotoxic T-lymphocyte induction and antitumor activity. J Virol 73:7877–7881Google Scholar