title: Immunohistochemical mapping and transcript expression of the GPA2/GPB5 receptor in tissues of the adult mosquito, Aedes aegypti.
abstract: GPA2/GPB5 is a glycoprotein hormone found in most bilateral metazoans including the mosquito, Aedes aegypti. To elucidate physiological roles and functions of GPA2/GPB5, we aim to identify prospective target tissues by examining the tissue- and sex-specific expression profile of its receptor, the leucine-rich repeat-containing G protein-coupled receptor 1 (LGR1) in the adult mosquito. Western analyses using a heterologous system with CHO-K1 cells, transiently expressing A. aegypti LGR1, yielded a 112-kDa monomeric band and high-molecular weight multimers, which associated with membrane-protein fractions. Moreover, immunoblot analyses on protein isolated from HEK 293 T cells stably expressing a fusion construct of A. aegypti LGR1-EGFP (LGR1: 105 kDa+EGFP: 27 kDa) yielded a band with a measured molecular weight of 139 kDa that also associated with membrane-protein fractions and upon deglycosylation, migrated as a lower molecular weight band of 132 kDa. Immunocytochemical analysis of HEK 293 T cells stably expressing this fusion construct confirmed EGFP fluorescence and LGR1-like immunoreactivity colocalized primarily to the plasma membrane. Immunohistochemical mapping in adult mosquitoes revealed LGR1-like immunoreactivity is widespread in the alimentary canal. Importantly, LGR1-like immunoreactivity localizes specifically to basolateral regions of epithelia and, in some regions, appeared as punctate intracellular staining, which together indicates a potential role in feeding and/or hydromineral balance. LGR1 transcript expression was also detected in gut regions that exhibited strong LGR1-like immunoreactivity. Interestingly, LGR1 transcript expression and strong LGR1-like immunoreactivity was also identified in reproductive tissues including the testes and ovaries, which together suggests a potential role linked to spermatogenesis and oogenesis in male and female mosquitoes, respectively.
authors.affiliation: Department of Biology, York University, Lumbers Building Room 221, 4700 Keele Street, Toronto, ON, M3J 1P3, Canada.
authors.forename: David A
authors.initials: DA
authors.lastname: Rocco
authors.affiliation: Department of Biology, York University, Lumbers Building Room 221, 4700 Keele Street, Toronto, ON, M3J 1P3, Canada.
authors.forename: Doo Hyun
authors.initials: DH
authors.lastname: Kim
authors.affiliation: Department of Biology, York University, Lumbers Building Room 221, 4700 Keele Street, Toronto, ON, M3J 1P3, Canada. paluzzi@yorku.ca.
authors.forename: Jean-Paul V
authors.initials: JV
authors.lastname: Paluzzi
mesh_terms: D000330:Aedes / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D020869:Gene Expression Profiling; D057809:HEK293 Cells; D006801:Humans; D007150:Immunohistochemistry; D019476:Insect Proteins / Q000235:genetics* / Q000378:metabolism; D009928:Organ Specificity; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism; D014162:Transfection
chemical_list: D019476:Insect Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface
keywords: Aedes aegypti; GPA2/GPB5; Glycoprotein hormone; LGR1; Leucine-rich repeat-containing G protein-coupled receptor 1; Mosquito