Alpha-Gal Syndrome: Involvement of Amblyomma americanum alpha-D-Galactosidase and beta-1,4 Galactosyltransferase Enzymes in alpha-Gal Metabolism

Alpha-Gal Syndrome (AGS) is an IgE-mediated delayed-type hypersensitivity reaction to the oligosaccharide galactose-alpha-1, 3-galactose (alpha-gal) injected into humans from the lone-star tick (Amblyomma americanum) bite. Indeed, alpha-gal is discovered in salivary glands of lone-star tick; however, the tick's specific intrinsic factors involved in endogenous alpha-gal production and presentation to host during hematophagy are poorly understood. This study aimed to investigate the functional role of two tick enzymes, alpha-D-galactosidase (ADGal) and beta-1,4 galactosyltransferases (beta-1,4GalT), in endogenous alpha-gal production, carbohydrate metabolism, and N-glycan profile in lone-star tick. The ADGal enzyme cleaves terminal alpha-galactose moieties from glycoproteins and glycolipids, whereas beta-1,4GalT transfers alpha-galactose to a beta 1,4 terminal linkage acceptor sugars-GlcNAc, Glc, and Xyl-in various processes of glycoconjugate synthesis. An RNA interference approach was utilized to silence ADGal and beta-1,4GalT in Am. americanum to examine their function in alpha-gal metabolism in tick and AGS onset. Silencing of ADGal led to the significant downregulation of genes involved in galactose metabolism and transport in Am. americanum. Immunoblot and N-glycan analysis of the Am. americanum salivary glands showed a significant reduction in alpha-gal levels in silenced tissues. However, there was no significant difference in the level of alpha-gal in beta-1,4GalT-silenced tick salivary glands. A basophil-activation test showed a decrease in the frequency of activated basophil by ADGal-silenced salivary glands. These results provide an insight into the roles of ADGal and beta-1,4GalT in alpha-gal production and presentation in ticks and the probable involvement in the onset of AGS.