title: Distinctive subcellular inhibition of cytokine-induced SRC by salubrinal and fluid flow.
abstract: A non-receptor protein kinase Src plays a crucial role in fundamental cell functions such as proliferation, migration, and differentiation. While inhibition of Src is reported to contribute to chondrocyte homeostasis, its regulation at a subcellular level by chemical inhibitors and mechanical stimulation has not been fully understood. In response to inflammatory cytokines and stress to the endoplasmic reticulum (ER) that increase proteolytic activities in chondrocytes, we addressed two questions: Do cytokines such as interleukin 1 beta (IL1β) and tumor necrosis factor alpha (TNFα) induce location-dependent Src activation? Can cytokine-induced Src activation be suppressed by chemically alleviating ER stress or by applying fluid flow? Using live cell imaging with two Src biosensors (i.e., cytosolic, and plasma membrane-bound biosensors) for a fluorescence resonance energy transfer (FRET) technique, we determined cytosolic Src activity as well as membrane-bound Src activity in C28/I2 human chondrocytes. In response to TNFα and IL1β, both cytosolic and plasma membrane-bound Src proteins were activated, but activation in the cytosol occurred earlier than that in the plasma membrane. Treatment with salubrinal or guanabenz, two chemical agents that attenuate ER stress, significantly decreased cytokine-induced Src activities in the cytosol, but not in the plasma membrane. In contrast, fluid flow reduced Src activities in the plasma membrane, but not in the cytosol. Collectively, the results demonstrate that Src activity is differentially regulated by salubrinal/guanabenz and fluid flow in the cytosol and plasma membrane.
authors.affiliation: Department of Biomedical Engineering, Indiana University-Purdue University Indianapolis, Indianapolis, Indiana, United States of America.
authors.forename: Qiaoqiao
authors.initials: Q
authors.lastname: Wan
authors.affiliation: Department of Orthopedics, Second Clinical Hospital of Harbin Medical University, Harbin, China.
authors.forename: Wenxiao
authors.initials: W
authors.lastname: Xu
authors.affiliation: Department of Orthopedics, Second Clinical Hospital of Harbin Medical University, Harbin, China.
authors.forename: Jing-long
authors.initials: JL
authors.lastname: Yan
authors.affiliation: Department of Biomedical Engineering, Indiana University-Purdue University Indianapolis, Indianapolis, Indiana, United States of America; Department of Anatomy and Cell Biology, Indiana University School of Medicine, Indianapolis, Indiana, United States of America.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Yokota
authors.affiliation: Department of Biomedical Engineering, Indiana University-Purdue University Indianapolis, Indianapolis, Indiana, United States of America.
authors.forename: Sungsoo
authors.initials: S
authors.lastname: Na
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology / Q000276:immunology; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000201:enzymology* / Q000276:immunology; D002934:Cinnamates / Q000494:pharmacology*; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology / Q000276:immunology; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D053583:Interleukin-1beta / Q000276:immunology*; D012212:Rheology; D013314:Stress, Mechanical; D013890:Thiourea / Q000031:analogs & derivatives* / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology*; D019061:src-Family Kinases / Q000037:antagonists & inhibitors* / Q000276:immunology*
chemical_list: D002934:Cinnamates; D053583:Interleukin-1beta; D014409:Tumor Necrosis Factor-alpha; C496827:salubrinal; D019061:src-Family Kinases; D013890:Thiourea