title: Dynamic regulation of aquaglyceroporin expression in erythrocyte cultures from cold- and warm-acclimated Cope's gray treefrog, Hyla chrysoscelis.
abstract: Cope's gray treefrog, Hyla chrysoscelis,is a freeze-tolerant anuran which accumulates and distributes glycerol as a cryoprotectant before freezing. We hypothesize that HC-3, an aquaglyceroporin member of the MIP family of water pores, may play an important role in the process of freeze tolerance by mediating transmembrane passage of glycerol and water during cold-acclimation. The objectives of this study were two-fold: to examine HC-3 protein abundance and cellular localization in erythrocytes from cold- and warm-acclimated frogs and to develop and characterize an erythrocyte cell culture system for examining HC-3 gene regulation. Compared with warm-acclimated frogs, erythrocytes from cold-acclimated frogs had higher HC-3 protein expression and enhanced plasma membrane localization. Furthermore, erythrocytes from cold- and warm-acclimated frogs maintained in culture at 4 and 20°C exhibited time- and temperature-dependent regulation of HC-3 expression and an increase in the abundance of high molecular weight immunoreactive species within 24 hr of culture at 20°C. Deglycosylation of erythrocyte proteins resulted in the disappearance of the high molecular weight species, indicating that HC-3 is post-translationally modified by N-linked glycosylation. Erythrocytes cultured in media containing glycerol also showed an increased abundance of the high molecular weight bands and enhanced plasma membrane localization of HC-3, suggesting a role for glycerol in regulating HC-3 subcellular trafficking. Thus, the development of this erythrocyte cell culture system from H. chrysoscelis opened an opportunity to study the properties of cells with changing expression of an aquaglyceroporin, HC-3, and to explore the factors regulating that expression.
authors.affiliation: Department of Biology, University of Dayton, Dayton, Ohio 45463, USA.
authors.forename: Venkateshwar
authors.initials: V
authors.lastname: Mutyam
authors.forename: Matthew V
authors.initials: MV
authors.lastname: Puccetti
authors.forename: James
authors.initials: J
authors.lastname: Frisbie
authors.forename: David L
authors.initials: DL
authors.lastname: Goldstein
authors.forename: Carissa M
authors.initials: CM
authors.lastname: Krane
mesh_terms: D000064:Acclimatization / Q000502:physiology*; D000818:Animals; D001001:Anura / Q000502:physiology*; D051397:Aquaglyceroporins / Q000032:analysis / Q000378:metabolism*; D015153:Blotting, Western; D002470:Cell Survival; D002478:Cells, Cultured; D003080:Cold Temperature; D004912:Erythrocytes / Q000737:chemistry / Q000378:metabolism*; D006358:Hot Temperature; D007150:Immunohistochemistry; D008297:Male
chemical_list: D051397:Aquaglyceroporins