Hydrodynamic DNA vaccination for induction of mouse monoclonal antibodies specifically reacting with N-acetylgalactosaminide α2,6-sialyltransferase II (ST6GalNAc II)
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1
Palacky University in Olomouc, Faculty of Medicine, Department of Immunology, Czechia
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2
University of Alabama at Birmingham, Department of Microbiology, United States
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3
Veterinary Research Institute, Department of Virology and Diagnostics, Czechia
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4
Juntendo University, Faculty of Medicine, Japan
IgA nephropathy (IgAN) is associated with galactose (Gal) deficiency of some IgA1 hinge-region (HR) O-glycans. Gal-deficient IgA1 plays a key role in IgAN, as such molecules are recognized by anti-glycan antibodies, form immune complexes that deposit in the glomerular mesangium and induce glomerular injury. O-glycosylation of IgA1 is initiated in Golgi apparatus of IgA1-producing cells by attachment of N-acetylgalactosamine (GalNAc) to some Ser/Thr residues in the HR, followed by addition of Gal to the GalNAc. Sialic acid (SA) could be attached to Gal by an α2,3-sialyltransferase and/or to GalNAc by α2,6-sialyltransferase II (ST6GalNAc II). Premature SA attachment to GalNAc would prevent addition of Gal, thus resulting in Gal deficiency.
The goal of our study is to determine whether overexpression of ST6GalNAc II, described in IgA1-producing cells from IgAN patients, is associated with abnormal localization of the enzyme, i.e., a shift from trans-Golgi toward cis-Golgi apparatus, and thus contributes to production of Gal-deficient IgA1. We developed ST6GalNAc II-specific monoclonal antibody for fluorescence-microscopy analysis of ST6GalNAc II intracellular localization. cDNA of human ST6GalNAc II was cloned into DNA vaccination plasmid and hydrodynamically administered to mice. Recombinant ST6GalNAc II protein was used for screening of ST6GalNAc II-specific antibodies from sera of DNA-immunized mice and monoclonal antibodies secreted by hybridoma cells. Analyses by fluorescence microscopy revealed that IgA1-producing cells from IgAN patients have increased fluorescence signal for ST6GalNAc II in trans-Golgi and a partial shift toward the cis-Golgi compared to IgA1-producing cells from healthy controls.
Supported by P302/10/1055, LH11046, DK078244, DK082753, and DK083663.
Keywords:
DNA vaccination,
Glycosyltransferases,
ST6GalNAc II,
IgA1,
IgA nephropathy
Conference:
15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013.
Presentation Type:
Abstract
Topic:
Adaptive Immunity
Citation:
Stuchlova Horynova
M,
Kasperova
A,
Smrzova
Z,
Weigl
E,
Suzuki
H,
Novak
J and
Raska
M
(2013). Hydrodynamic DNA vaccination for induction of mouse monoclonal antibodies specifically reacting with N-acetylgalactosaminide α2,6-sialyltransferase II (ST6GalNAc II).
Front. Immunol.
Conference Abstract:
15th International Congress of Immunology (ICI).
doi: 10.3389/conf.fimmu.2013.02.00320
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Received:
15 Mar 2013;
Published Online:
22 Aug 2013.
*
Correspondence:
Mrs. Milada Stuchlova Horynova, Palacky University in Olomouc, Faculty of Medicine, Department of Immunology, Olomouc, Czechia, stuchlova.horynova@seznam.cz