| Autor: |
Julia E. Hölper, Finn Grey, John Kenneth Baillie, Tim Regan, Nicholas J. Parkinson, Dirk Höper, Thiprampai Thamamongood, Martin Schwemmle, Katrin Pannhorst, Lisa Wendt, Thomas C. Mettenleiter, Barbara G. Klupp |
| Jazyk: |
angličtina |
| Rok vydání: |
2021 |
| Předmět: |
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| Zdroj: |
Viruses, Vol 13, Iss 8, p 1574 (2021) |
| Druh dokumentu: |
article |
| ISSN: |
1999-4915 |
| DOI: |
10.3390/v13081574 |
| Popis: |
Herpesviruses are large DNA viruses, which encode up to 300 different proteins including enzymes enabling efficient replication. Nevertheless, they depend on a multitude of host cell proteins for successful propagation. To uncover cellular host factors important for replication of pseudorabies virus (PrV), an alphaherpesvirus of swine, we performed an unbiased genome-wide CRISPR/Cas9 forward screen. To this end, a porcine CRISPR-knockout sgRNA library (SsCRISPRko.v1) targeting 20,598 genes was generated and used to transduce porcine kidney cells. Cells were then infected with either wildtype PrV (PrV-Ka) or a PrV mutant (PrV-gD–Pass) lacking the receptor-binding protein gD, which regained infectivity after serial passaging in cell culture. While no cells survived infection with PrV-Ka, resistant cell colonies were observed after infection with PrV-gD–Pass. In these cells, sphingomyelin synthase 1 (SMS1) was identified as the top hit candidate. Infection efficiency was reduced by up to 90% for PrV-gD–Pass in rabbit RK13-sgms1KO cells compared to wildtype cells accompanied by lower viral progeny titers. Exogenous expression of SMS1 partly reverted the entry defect of PrV-gD–Pass. In contrast, infectivity of PrV-Ka was reduced by 50% on the knockout cells, which could not be restored by exogenous expression of SMS1. These data suggest that SMS1 plays a pivotal role for PrV infection, when the gD-mediated entry pathway is blocked. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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