Evaluating large spontaneous deletions in a bovine cell line selected for bovine viral diarrhea virus resistance

Authors

Aspen M. Workman, USDA ARS Roman L. Hruska U.S. Meat Animal Research CenterFollow
Michael P. Heaton, USDA, ARS, U.S. Meat Animal Research CenterFollow
Dennis A. Webster, Recombinetics, Inc.Follow
Gregory P. Harhay, USDA ARS Roman L. Hruska U.S. Meat Animal Research CenterFollow
Theodore S. Kalbfleisch, University of Kentucky, LexingtonFollow
Timothy P.L. Smith, USDA ARS Roman L. Hruska U.S. Meat Animal Research CenterFollow
Shollie M. Falkenberg, USDA ARS National Animal Disease CenterFollow
Daniel F. Carlson, Recombinetics, Inc.Follow
Tad S. Sonstegard, AcceligenFollow

ORCID IDs

Aspen M. Workman https://orcid.org/0000-0003-2852-3519

Michael P. Heaton https://orcid.org/0000-0003-1386-1208

Timothy P. L. Smith https://orcid.org/0000-0003-1611-6828

Date of this Version

11-1-2021

Document Type

Article

Citation

Workman, A.M.; Heaton, M.P.;Webster, D.A.; Harhay, G.P.; Kalbfleisch, T.S.; Smith, T.P.L.; Falkenberg, S.M.; Carlson, D.F.; Sonstegard, T.S. Evaluating Large Spontaneous Deletions in a Bovine Cell Line Selected for Bovine Viral Diarrhea Virus Resistance. Viruses 2021, 13, 2147. https://doi.org/ 10.3390/v13112147

Comments

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license

Abstract

Bovine viral diarrhea virus’s (BVDV) entry into bovine cells involves attachment of virions to cellular receptors, internalization, and pH-dependent fusion with endosomal membranes. The primary host receptor for BVDV is CD46; however, the complete set of host factors required for virus entry is unknown. The Madin-Darby bovine kidney (MDBK) cell line is susceptible to BVDV infection, while a derivative cell line (CRIB) is resistant at the level of virus entry. We performed complete genome sequencing of each to identify genomic variation underlying the resistant phenotype with the aim of identifying host factors essential for BVDV entry. Three large compound deletions in the BVDV-resistant CRIB cell line were identified and predicted to disrupt the function or expression of the genes PTPN12, GRID2, and RABGAP1L. However, CRISPR/Cas9 mediated knockout of these genes, individually or in combination, in the parental MDBK cell line did not impact virus entry or replication. Therefore, resistance to BVDV in the CRIB cell line is not due to the apparent spontaneous loss of PTPN12, GRID2, or RABGAP1L gene function. Identifying the functional cause of BVDV resistance in the CRIB cell line may require more detailed comparisons of the genomes and epigenomes.