title: Dissection of Thrombospondin-4 Domains Involved in Intracellular Adaptive Endoplasmic Reticulum Stress-Responsive Signaling.
abstract: Thrombospondins are a family of stress-inducible secreted glycoproteins that underlie tissue remodeling. We recently reported that thrombospondin-4 (Thbs4) has a critical intracellular function, regulating the adaptive endoplasmic reticulum (ER) stress pathway through activating transcription factor 6α (Atf6α). In the present study, we dissected the domains of Thbs4 that mediate interactions with ER proteins, such as BiP (Grp78) and Atf6α, and the domains mediating activation of the ER stress response. Functionally, Thbs4 localized to the ER and post-ER vesicles and was actively secreted from cardiomyocytes, as were the type III repeat (T3R) and TSP-C domains, while the LamG domain localized to the Golgi apparatus. We also mutated the major calcium-binding motifs within the T3R domain of full-length Thbs4, causing ER retention and secretion blockade. The T3R and TSP-C domains as well as wild-type Thbs4 and the calcium-binding mutant interacted with Atf6α, induced an adaptive ER stress response, and caused expansion of intracellular vesicles. In contrast, overexpression of a related secreted oligomeric glycoprotein, Nell2, which lacks only the T3R and TSP-C domains, did not cause these effects. Finally, deletion of Atf6α abrogated Thbs4-induced vesicular expansion. Taken together, these data identify the critical intracellular functional domains of Thbs4, which was formerly thought to have only extracellular functions.
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Brody
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: Tobias G
authors.initials: TG
authors.lastname: Schips
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: Davy
authors.initials: D
authors.lastname: Vanhoutte
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: Onur
authors.initials: O
authors.lastname: Kanisicak
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA Howard Hughes Medical Institute, Cincinnati, Ohio, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Karch
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: Bryan D
authors.initials: BD
authors.lastname: Maliken
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: N Scott
authors.initials: NS
authors.lastname: Blair
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA Howard Hughes Medical Institute, Cincinnati, Ohio, USA.
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Sargent
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA.
authors.forename: Vikram
authors.initials: V
authors.lastname: Prasad
authors.affiliation: Department of Pediatrics, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio, USA Howard Hughes Medical Institute, Cincinnati, Ohio, USA jeff.molkentin@cchmc.org.
authors.forename: Jeffery D
authors.initials: JD
authors.lastname: Molkentin
mesh_terms: D051702:Activating Transcription Factor 6 / Q000378:metabolism; D000818:Animals; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D059865:Endoplasmic Reticulum Stress / Q000235:genetics* / Q000502:physiology*; D006023:Glycoproteins / Q000235:genetics; D006056:Golgi Apparatus / Q000378:metabolism*; D006360:Heat-Shock Proteins; D051379:Mice; D008822:Mice, Transgenic; D032383:Myocytes, Cardiac / Q000378:metabolism / Q000648:ultrastructure; D021381:Protein Transport / Q000235:genetics; D051381:Rats; D015398:Signal Transduction / Q000235:genetics; D019699:Thrombospondins / Q000235:genetics / Q000378:metabolism*
chemical_list: D051702:Activating Transcription Factor 6; D000091342:Endoplasmic Reticulum Chaperone BiP; D006023:Glycoproteins; D006360:Heat-Shock Proteins; C000717248:Hspa5 protein, mouse; D019699:Thrombospondins; C101864:thrombospondin 4