Journal article
Expression of bovine non-classical major histocompatibility complex class I proteins in mouse P815 and human K562 cells
Research in veterinary science, Vol.107, pp.161-170
08/2016
Handle:
https://hdl.handle.net/2376/117024
PMID: 27473990
Abstract
Major histocompatibility complex class I (MHC-I) proteins can be expressed as cell surface or secreted proteins. To investigate whether bovine non-classical MHC-I proteins are expressed as cell surface or secreted proteins, and to assess the reactivity pattern of monoclonal antibodies with non-classical MHC-I isoforms, we expressed the MHC proteins in murine P815 and human K562 (MHC-I deficient) cells. Following antibiotic selection, stably transfected cell lines were stained with H1A or W6/32 antibodies to detect expression of the MHC-I proteins by flow cytometry. Two non-classical proteins (BoLA-NC1*00501 and BoLA-NC3*00101) were expressed on the cell surface in both cell lines. Surprisingly, the BoLA-NC4*00201 protein was expressed on the cell membrane of human K562 but not mouse P815 cells. Two non-classical proteins (BoLA-NC1*00401, which lacks a transmembrane domain, and BoLA-NC2*00102) did not exhibit cell surface expression. Nevertheless, Western blot analyses demonstrated expression of the MHC-I heavy chain in all transfected cell lines. Ammonium-sulfate precipitation of proteins from culture supernatants showed that BoLA-NC1*00401 was secreted and that all surface expressed proteins where shed from the cell membrane by the transfected cells. Interestingly, the surface expressed MHC-I proteins were present in culture supernatants at a much higher concentration than BoLA-NC1*00401. This comprehensive study shows that bovine non-classical MHC-I proteins BoLA-NC1*00501, BoLA-NC3*00101, and BoLA-NC4*00201 are expressed as surface isoforms with the latter reaching the cell membrane only in K562 cells. Furthermore, it demonstrated that BoLA-NC1*00401 is a secreted isoform and that significant quantities of membrane associated MHC-I proteins can be shed from the cell membrane.
•Bovine non-classical MHC-I proteins can be either membrane associated or secreted.•BoLA-NC1*00501, -NC3*00101 and -NC4*00201 were membrane associated proteins.•The BoLA-NC1*00401 splice variant was expressed as a secreted protein.•BoLA-NC2*00102 remained intracellular and may require a specific peptide antigen.
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Details
- Title
- Expression of bovine non-classical major histocompatibility complex class I proteins in mouse P815 and human K562 cells
- Creators
- Parveen Parasar - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USAAmanda Wilhelm - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USAHeloisa M Rutigliano - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USAAaron J Thomas - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USALihong Teng - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USABi Shi - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USAWilliam C Davis - Department of Veterinary Microbiology and Pathology, P.O. Box 647040, Washington State University, Pullman, WA, USACarlos E Suarez - USDA-ARS Animal Disease Research Unit, P.O. Box 646630, Washington State University, Pullman, WA, USADaniel D New - Department of Veterinary Microbiology and Pathology, P.O. Box 647040, Washington State University, Pullman, WA, USAKenneth L White - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USAChristopher J Davies - Department of Animal, Dairy and Veterinary Sciences, 4815 Old Main Hill, Utah State University, Logan, UT, USA
- Publication Details
- Research in veterinary science, Vol.107, pp.161-170
- Academic Unit
- Veterinary Microbiology and Pathology, Department of
- Publisher
- Elsevier Ltd
- Identifiers
- 99900547727001842
- Language
- English
- Resource Type
- Journal article