title: Keratinocyte secretion of cyclophilin B via the constitutive pathway is regulated through its cyclosporin-binding site.
abstract: Cyclophilin B (CypB) is an endoplasmic reticulum (ER)-resident member of the cyclophilin family of proteins that bind cyclosporin A (CsA). We report that as in other cell types, CypB trafficked from the ER and was secreted by keratinocytes into the media in response to CsA. Concentrations as low as 1 pM of CsA induced secretion of CypB. Using brefeldin A, we showed that CypB is secreted from keratinocytes via the constitutive secretory pathway. We defined that substitution of tryptophan residue 128 in the CsA-binding site of CypB with alanine resulted in dissociation of CypB(W128A)-green fluorescent protein (GFP) from the ER. Photobleaching studies revealed a significant reduction in the diffusible mobility of CypB(W128A)-GFP compared with CypB(WT)-GFP, consistent with redistribution of CypB(W128A)-GFP into secretory vesicles disconnected from the ER/Golgi network. Furthermore, CsA significantly decreased the mobility of CypB(WT)-GFP but not CypB(W128A)-GFP. These studies demonstrate that therapeutically relevant concentrations of CsA regulate secretion of CypB by keratinocytes, and that a key residue within the CsA-binding site of CypB controls retention of CypB within the ER and regulates entry into the secretory pathway. As keratinocytes express CypB receptors (CD147) and CypB exhibits chemotactic properties, these data have implications for the therapeutic effects of CsA in inflammatory skin disease.
authors.affiliation: Institute of Cellular Medicine, Medical School, Newcastle University, Newcastle Upon Tyne, UK.
authors.forename: Paula
authors.initials: P
authors.lastname: Fearon
authors.forename: Ann A
authors.initials: AA
authors.lastname: Lonsdale-Eccles
authors.forename: O Kehinde
authors.initials: OK
authors.lastname: Ross
authors.forename: Carole
authors.initials: C
authors.lastname: Todd
authors.forename: Aparna
authors.initials: A
authors.lastname: Sinha
authors.forename: Fabrice
authors.initials: F
authors.lastname: Allain
authors.forename: Nick J
authors.initials: NJ
authors.lastname: Reynolds
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D021983:Cyclophilins / Q000378:metabolism*; D016572:Cyclosporine / Q000494:pharmacology*; D003879:Dermatologic Agents / Q000494:pharmacology*; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology; D006056:Golgi Apparatus / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D015603:Keratinocytes / Q000187:drug effects* / Q000201:enzymology*; D022142:Secretory Vesicles / Q000187:drug effects / Q000201:enzymology
chemical_list: D003879:Dermatologic Agents; C081786:cyclophilin B; D016572:Cyclosporine; D021983:Cyclophilins