title: Cellular Factor XIIIA Transglutaminase Localizes in Caveolae and Regulates Caveolin-1 Phosphorylation, Homo-oligomerization and c-Src Signaling in Osteoblasts.
abstract: Transglutaminases (TGs) are a family of widely distributed enzymes that catalyze protein crosslinking by forming a covalent isopeptide bond between the substrate proteins. We have shown that MC3T3-E1 osteoblasts express Factor XIII-A (FXIII-A), and that the extracellular crosslinking activity of FXIII-A is involved in regulating matrix secretion and deposition. In this study, we have investigated the localization and potential role of intracellular FXIII-A. Conventional immunofluorescence microscopy and TIRF microscopy analyses showed that FXIII-A co-localizes with caveolin-1 in specialized membrane structures, caveolae, in differentiating osteoblasts. The caveolae-disrupting agent methyl-β-cyclodextrin abolished FXIII-A staining and co-localization with caveolin-1 from the osteoblast plasma membrane. The presence of FXIII-A in caveolae was confirmed by preparing caveolae-enriched cellular fractions using sucrose density gradient ultracentrifugation followed by western blotting. Despite this association of FXIII-A with caveolae, there was no detectable transglutaminase activity in caveolae, as measured by monodansylcadaverine incorporation. TG inhibitor NC9--which can alter TG enzyme conformation--localized to caveolae and displaced FXIII-A from these structures when added to the osteoblast cultures. The decreased FXIII-A levels in caveolae after NC9 treatment increased c-Src activation, which resulted in caveolin-1 phosphorylation, homo-oligomerization and Akt phosphorylation, suggesting cellular FXIII-A has a role in regulating c-Src signaling in osteoblasts.
authors.affiliation: Division of Biomedical Sciences, Faculty of Dentistry, McGill University, Montreal, QC, Canada. (SW, MTK)
authors.forename: Shuai
authors.initials: S
authors.lastname: Wang
authors.affiliation: Division of Experimental Medicine, Department of Medicine, Faculty of Medicine, McGill University, Montreal, QC, Canada (MTK)|Division of Biomedical Sciences, Faculty of Dentistry, McGill University, Montreal, QC, Canada. (SW, MTK)
authors.forename: Mari T
authors.initials: MT
authors.lastname: Kaartinen
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001704:Biopolymers / Q000378:metabolism*; D021941:Caveolae / Q000201:enzymology*; D051242:Caveolin 1 / Q000378:metabolism*; D026122:Factor XIIIa / Q000378:metabolism*; D051379:Mice; D010006:Osteoblasts / Q000201:enzymology / Q000378:metabolism*; D010766:Phosphorylation; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D015398:Signal Transduction; D011503:Transglutaminases / Q000378:metabolism*
chemical_list: D001704:Biopolymers; D051242:Caveolin 1; D026122:Factor XIIIa; D011503:Transglutaminases; D016392:Proto-Oncogene Proteins pp60(c-src)
keywords: Factor XIII-A transglutaminase; c-Src kinase; caveolae; caveolin-1; osteoblast differentiation