title: Transglutaminase 2-dependent deamidation of glyceraldehyde-3-phosphate dehydrogenase promotes trophoblastic cell fusion.
abstract: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional protein as well as a classic glycolytic enzyme, and its pleiotropic functions are achieved by various post-translational modifications and the resulting translocations to intracellular compartments. In the present study, GAPDH in the plasma membrane of BeWo choriocarcinoma cells displayed a striking acidic shift in two-dimensional electrophoresis after cell-cell fusion induction by forskolin. This post-translational modification was deamidation of multiple glutaminyl residues, as determined by molecular mass measurement and tandem mass spectrometry of acidic GAPDH isoforms. Transglutaminase (TG) inhibitors prevented this acidic shift and reduced cell fusion. Knockdown of the TG2 gene by short hairpin RNA reproduced these effects of TG inhibitors. Various GAPDH mutants with replacement of different numbers (one to seven) of Gln by Glu were expressed in BeWo cells. These deamidated mutants reversed the suppressive effect of wild-type GAPDH overexpression on cell fusion. Interestingly, the mutants accumulated in the plasma membrane, and this accumulation was increased according to the number of Gln/Glu substitutions. Considering that GAPDH binds F-actin via an electrostatic interaction and that the cytoskeleton is rearranged in trophoblastic cell fusion, TG2-dependent GAPDH deamidation was suggested to participate in actin cytoskeletal remodeling.
authors.affiliation: From the Department of Molecular Medicine, Osaka Medical Center and Research Institute for Maternal and Child Health, 840 Murodo-cho, Izumi, Osaka 594-1101, Japan and.
authors.forename: Kaori
authors.initials: K
authors.lastname: Iwai
authors.forename: Yukinao
authors.initials: Y
authors.lastname: Shibukawa
authors.forename: Natsuko
authors.initials: N
authors.lastname: Yamazaki
authors.forename: Yoshinao
authors.initials: Y
authors.lastname: Wada
mesh_terms: D000577:Amides / Q000378:metabolism*; D000595:Amino Acid Sequence; D002459:Cell Fusion; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D015726:Giant Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011503:Transglutaminases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014327:Trophoblasts / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*
chemical_list: D000577:Amides; D004791:Enzyme Inhibitors; D050505:Mutant Proteins; D005973:Glutamine; D018698:Glutamic Acid; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins
keywords: Cell Differentiation; Cell Fusion; Cytoskeleton; Glyceraldehyde-3-phosphate Dehydrogenase; Protein Deamidation; Transglutaminases; Trophoblast