ADAM17 Is an Essential Factor for the Infection of Bovine Cells with Pestiviruses.

Marianne Zaruba,Stefan Düsterhöft,Aspen M Workman,Kati Szakmary-Braendle,Till Rümenapf,Kerstin Seitz,Ole Frithjof Pietsch,Angelika Auer,Christiane Riedel,Marlene Mötz,Hann-Wei Chen

doi:10.3390/v14020381

Abstract

The entry of BVDV into bovine cells was studied using CRIB cells (cells resistant to infection with bovine viral diarrhea virus [BVDV]) that have evolved from MDBK cells by a spontaneous loss of susceptibility to BVDV. Recently, larger genetic deletions were reported but no correlation of the affected genes and the resistance to BVDV infection could be established. The metalloprotease ADAM17 was reported as an essential attachment factor for the related classical swine fever virus (CSFV). To assess whether ADAM17 might be involved in the resistance of CRIB-1 cells to pestiviruses, we analyzed its expression in CRIB-1 and MDBK cells. While ADAM17 protein was detectable in MBDK cells, it was absent from CRIB-1 cells. No functional full-length ADAM17 mRNA could be detected in CRIB cells and genetic analysis revealed the presence of two defective alleles. Transcomplementation of functional ADAM17 derived from MDBK cells in CRIB-1 cells resulted in a nearly complete reversion of their resistance to pestiviral infection. Our results demonstrate that ADAM17 is a key cellular factor for the pestivirus resistance of CRIB-1 cells and establishes its essential role for a broader range of pestiviruses.

Highlights

Pestiviruses are small, enveloped viruses with a single stranded, positive sense RNA genome of about 12.3 kb
ADAM17 Alleles Are Disrupted in CRIB-1 Cells
We put forward three lines of evidence that ADAM17 is the missing factor causing the resistance of CRIB-1 cells to pestivirus infection

Summary

Introduction

Pestiviruses (family Flaviviridae) are small, enveloped viruses with a single stranded, positive sense RNA genome of about 12.3 kb. Pestiviral particles harbor three glycoproteins in their envelope, Erns , E1 and E2. The glycoproteins E1 and E2 are usually present as a heterodimer in the virus particle [1,2], and E2 is responsible for receptor binding and together with Erns a target of neutralizing antibodies [1,3,4]. Several cell surface proteins have been identified as mediators of pestivirus entry. For BVDV and atypical porcine pestivirus (APPV), CD46 was discovered as a receptor molecule that interacts with E2 [12–14]. After binding to a receptor molecule, pestivirus entry proceeds, as experimentally shown for BVDV and CSFV, by clathrin-mediated endocytosis [19–22] that depends on the presence of cholesterol and functional dynamin. After clathrin-mediated uptake, CSFV particles colocalize with RAB5-(Ras-like GTPases) and RAB7-positive endosomes [21], indicating that pestiviral fusion takes place in the low pH compartment of late endosomes or lysosomes

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