title: TDAG51 mediates epithelial-to-mesenchymal transition in human proximal tubular epithelium.
abstract: Epithelial-to-mesenchymal transition (EMT) contributes to renal fibrosis in chronic kidney disease. Endoplasmic reticulum (ER) stress, a feature of many forms of kidney disease, results from the accumulation of misfolded proteins in the ER and leads to the unfolded protein response (UPR). We hypothesized that ER stress mediates EMT in human renal proximal tubules. ER stress is induced by a variety of stressors differing in their mechanism of action, including tunicamycin, thapsigargin, and the calcineurin inhibitor cyclosporine A. These ER stressors increased the UPR markers GRP78, GRP94, and phospho-eIF2α in human proximal tubular cells. Thapsigargin and cyclosporine A also increased cytosolic Ca(2+) concentration and T cell death-associated gene 51 (TDAG51) expression, whereas tunicamycin did not. Thapsigargin was also shown to increase levels of active transforming growth factor (TGF)-β1 in the media of cultured human proximal tubular cells. Thapsigargin induced cytoskeletal rearrangement, β-catenin nuclear translocation, and α-smooth muscle actin and vinculin expression in proximal tubular cells, indicating an EMT response. Subconfluent primary human proximal tubular cells were induced to undergo EMT by TGF-β1 treatment. In contrast, tunicamycin treatment did not produce an EMT response. Plasmid-mediated overexpression of TDAG51 resulted in cell shape change and β-catenin nuclear translocation. These results allowed us to develop a two-hit model of ER stress-induced EMT, where Ca(2+) dysregulation-mediated TDAG51 upregulation primes the cell for mesenchymal transformation via Wnt signaling and then TGF-β1 activation leads to a complete EMT response. Thus the release of Ca(2+) from ER stores mediates EMT in human proximal tubular epithelium via the induction of TDAG51.
authors.affiliation: Department of Medicine, Division of Nephrology, McMaster University and St. Joseph’s Healthcare Hamilton, Hamilton, Ontario, Canada.
authors.forename: Rachel E
authors.initials: RE
authors.lastname: Carlisle
authors.forename: Alana
authors.initials: A
authors.lastname: Heffernan
authors.forename: Elise
authors.initials: E
authors.lastname: Brimble
authors.forename: Limin
authors.initials: L
authors.lastname: Liu
authors.forename: Danielle
authors.initials: D
authors.lastname: Jerome
authors.forename: Celeste A
authors.initials: CA
authors.lastname: Collins
authors.forename: Zahraa
authors.initials: Z
authors.lastname: Mohammed-Ali
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Margetts
authors.forename: Richard C
authors.initials: RC
authors.lastname: Austin
authors.forename: Jeffrey G
authors.initials: JG
authors.lastname: Dickhout
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D048430:Cell Shape; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D004797:Enzyme-Linked Immunosorbent Assay; D004848:Epithelium / Q000378:metabolism*; D005355:Fibrosis; D006801:Humans; D007202:Indicators and Reagents; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D008648:Mesoderm / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000235:genetics; D013050:Spectrometry, Fluorescence; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000096:biosynthesis; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D051176:beta Catenin / Q000378:metabolism
chemical_list: D002614:Chelating Agents; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; C000717248:Hspa5 protein, mouse; D007202:Indicators and Reagents; C491923:PHLDA1 protein, human; D014157:Transcription Factors; D053773:Transforming Growth Factor beta1; D051176:beta Catenin; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D002118:Calcium