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Glycosylation of aquaporins in the freeze tolerant tree frog, Hyla chrysoscelis
Author(s) -
Alnajjar Khadijeh,
Pandey Ram Naresh,
Frisbie James,
Goldstein David L.
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.598.9
Subject(s) - aquaporin , glycosylation , pngase f , biology , biochemistry , extracellular , microbiology and biotechnology , chemistry , glycoprotein , glycan
HC‐1, ‐2, and ‐3 are aquaporins expressed in tissues of the freeze‐tolerant treefrog Hyla chrysoscelis . We hypothesize that the aquaporins are involved in osmoregulation during cold acclimation and freezing. Western blots reveal distinct protein bands at the nominal MW of the proteins (29 ‐ 31 kDa), and also at higher MW. We hypothesized that the high MW "smear" represents glycosylated forms of aquaporin involved in proper protein folding and localization to the membrane. The purpose of the present project was to investigate the magnitude of this glycosylation and its importance in quantifying aquaporin expression. We used Peptide N‐glycosidase (PNGase F), an enzyme that cleaves the oligosaccharides attached to extracellular asparagine residue, to examine N‐linked glycosylation of HC‐1, 2, and 3. In all tissues examined (skin, muscle, stomach, liver, blood, kidney, small and large intestine), addition of PNGase F decreased the intensity of signal at high MW and increased the intensity of the 29 ‐ 31 kDa band. We conclude that variation in aquaporin protein expression in treefrogs experiencing cold or freezing is associated with changing patterns of glycosylation. However, even with PNGase F, we continue to observe high MW bands in some tissues and conditions; further studies are exploring whether this represents an alternative form, O‐linked, glycosylation. Supported by NSF IOB‐0517301 and APS UGSRF.