Počet záznamů: 1  

Heterologous avian system for quantitative analysis of Syncytin-1 interaction with ASCT2 receptor

  1. 1.
    0543827 - ÚMG 2022 RIV GB eng J - Článek v odborném periodiku
    Štafl, Kryštof - Trávníček, Martin - Kučerová, Dana - Pecnová, Lubomíra - Krchlíková, Veronika - Gáliková, Eliška - Stepanets, Volodymyr - Hejnar, Jiří - Trejbalová, Kateřina
    Heterologous avian system for quantitative analysis of Syncytin-1 interaction with ASCT2 receptor.
    Retrovirology. Roč. 18, č. 1 (2021), č. článku 15. E-ISSN 1742-4690
    Grant CEP: GA ČR GA17-14356S
    Institucionální podpora: RVO:68378050
    Klíčová slova: Retroviral receptor * Envelope glycoprotein * Envelope-receptor interaction * Syncytin-1 * Cell-cell fusion * NanoLuc luciferase
    Obor OECD: Virology
    Impakt faktor: 3.768, rok: 2021
    Způsob publikování: Open access
    https://retrovirology.biomedcentral.com/articles/10.1186/s12977-021-00558-0

    Background Human Syncytin-1 is a placentally-expressed cell surface glycoprotein of retroviral origin. After interaction with ASCT2, its cellular receptor, Syncytin-1 triggers cell-cell fusion and formation of a multinuclear syncytiotrophoblast layer of the placenta. The ASCT2 receptor is a multi-spanning membrane protein containing a protruding extracellular part called region C, which has been suggested to be a retrovirus docking site. Precise identification of the interaction site between ASCT2 and Syncytin-1 is challenging due to the complex structure of ASCT2 protein and the background of endogenous ASCT2 gene in the mammalian genome. Chicken cells lack the endogenous background and, therefore, can be used to set up a system with surrogate expression of the ASCT2 receptor. Results We have established a retroviral heterologous chicken system for rapid and reliable assessment of ectopic human ASCT2 protein expression. Our dual-fluorescence system proved successful for large-scale screening of mutant ASCT2 proteins. Using this system, we demonstrated that progressive deletion of region C substantially decreased the amount of ASCT2 protein. In addition, we implemented quantitative assays to determine the interaction of ASCT2 with Syncytin-1 at multiple levels, which included binding of the soluble form of Syncytin-1 to ASCT2 on the cell surface and a luciferase-based assay to evaluate cell-cell fusions that were triggered by Syncytin-1. Finally, we restored the envelope function of Syncytin-1 in a replication-competent retrovirus and assessed the infection of chicken cells expressing human ASCT2 by chimeric Syncytin-1-enveloped virus. The results of the quantitative assays showed that deletion of the protruding region C did not abolish the interaction of ASCT2 with Syncytin-1. Conclusions We present here a heterologous chicken system for effective assessment of the expression of transmembrane ASCT2 protein and its interaction with Syncytin-1. The system profits from the absence of endogenous ASCT2 background and implements the quantitative assays to determine the ASCT2-Syncytin-1 interaction at several levels. Using this system, we demonstrated that the protruding region C was essential for ASCT2 protein expression, but surprisingly, not for the interaction with Syncytin-1 glycoprotein.
    Trvalý link: http://hdl.handle.net/11104/0320945

     
     
Počet záznamů: 1  

  Tyto stránky využívají soubory cookies, které usnadňují jejich prohlížení. Další informace o tom jak používáme cookies.