Apoptosis Induced by Duck Reovirus p10.8 Protein in Primary Duck Embryonated Fibroblast and Vero E6 Cells

Abstract

Muscovy duck reovirus (MDRV) is an important poultry pathogen that causes high morbidity and mortality in ducklings. The mechanisms by which viruses kill susceptible cells, and ultimately produce diseases, in Muscovy duck remain poorly understood. In this study, we focused on the biologic functions of the MDRV p10.8 protein in vitro. The p10.8 protein is a small protein of MDRV that is encoded by the first open reading frame of the S4 segment. In our study, the p10.8-encoding gene was individually cloned and expressed in bacterial and eukaryotic cells. The p10.8 protein had no potential transmembrane domain; it shared no sequence similarity to other known fusion-associated small transmembrane proteins encoded by the avian reovirus, Nelson Bay virus or baboon reovirus; and it did not show any syncytium formation activity. The p10.8 protein induced apoptosis when expressed by itself in transfected primary Muscovy duck embryonic fibroblasts or in Vero E6 cells. Four assays were used to analyze the apoptosis induced by p10.8: DNA ladder formation; terminal deoxynucleotidyl transferase dUTP nick end labeling; enzyme-linked immunosorbent assay detection of cytoplasmic histone-associated DNA fragments; and nuclear staining with propidium iodide. Two deletion products, p10.8delta1 (1-63aa; amino acid position) and p10.8delta2 (64-96aa), were constructed. Deletion analysis suggests that p10.8delta1 (1-63aa) is important in mediating p10.8-induced apoptosis because its deletion abolishes induction of apoptosis.