|Williams, John - ROSLIN INSTITUTE|
Submitted to: Animal Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 4, 2004
Publication Date: January 21, 2005
Citation: Szperka, M.E., Connor, E.E., Paape, M.J., Williams, J.L., Bannerman, D.D. 2005. Characterization of bovine FAS-associated death domain gene. Animal Genetics. 36(1):63-66. Interpretive Summary: In well-managed herds, almost 50% of all clinical cases of mastitis are caused by Gram-negative bacteria. All of these bacteria contain a highly pro-inflammatory molecule called endotoxin or lipopolysaccharide (LPS), which is shed from the bacterial surface during replication or death. LPS has been implicated in much of the injury/dysfunction associated with Gram-negative infection. Cellular activation by LPS enables the host organism to mount an innate immune response to the bacterial infection. In addition to activation, LPS induces tissue injury and cell death (apoptosis) that can result in scarring. In the setting of mastitis, this injury may contribute to decreased milk output. The present study sequenced, mapped, cloned, and functionally characterized bovine FADD, and demonstrated a role for bovine FADD in mediating LPS-induced injury to bovine cells. Further, this study demonstrated that in comparison to its human homologue, bovine FADD fails to downregulate LPS-induced pro-inflammatory signaling. This latter finding may explain the exquisite sensitivity of cattle to LPS-induced inflammation relative to other species.
Technical Abstract: The Fas-associated death domain (FADD) protein is an adapter/signaling molecule that has been shown to function in human cells to promote apoptosis and to inhibit NF-kB activation. Because of the critical role that apoptosis and NF-kB play in a variety of disease states, we mapped the bovine FADD gene, sequenced bovine FADD cDNA, and characterized its expression in endothelial cells. Sequencing of bovine FADD revealed approximately 65% and 58% amino acid sequence identity to its human and murine homologues, respectively. Bovine FADD was mapped to chromosome 29 by radiation hybrid mapping. In addition, the functionality of bovine FADD was studied. Expression of a bovine FADD dominant-negative construct blocked bacterial lipopolysaccharide (LPS)- and TNF-a-induced apoptosis in bovine endothelial cells consistent with previous studies of human FADD. In contrast to human FADD, elevated expression of bovine FADD had no effect on LPS- or TNF-a-induced upregulation of NF-kB-dependent gene products as assayed by E-selectin expression. Thus, while the role of FADD in mediating apoptosis is conserved across species, its role in regulating NF-kB-dependent gene expression is not.