title: Src signaling involvement in Japanese encephalitis virus-induced cytokine production in microglia.
abstract: Numerous studies have demonstrated that the disease pathogenesis of Japanese encephalitis involves cytokine-mediated bystander damage. The mechanisms involved in the regulation of Japanese encephalitis virus (JEV)-induced cytokine expression are not well defined but rely mainly on the tight regulation of transcription factor NF-κB. The Src-family tyrosine kinases participate in diversity of cellular signaling and have been demonstrated in JEV-infected cells. A direct link leading from Src activation to NF-κB activation in JEV-induced cytokine expression is incompletely understood. Here, we report that Src-related Ras/Raf/extracellular signal-regulated kinase (ERK) cascades participate in NF-κB activation and consequent tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β) expression in JEV-infected microglia. Central microglia were capable of producing TNF-α and IL-1β after JEV infection. However, JEV infection had a negligible effect on triggering TNF-α and IL-1β production by neurons and astrocytes. The expression of TNF-α and IL-1β caused by JEV was accompanied by increased Src phosphorylation, Ras membrane association, Raf serine-338 as well as tyrosine-340 phosphorylation, ERK phosphorylation, NF-κB DNA binding activity, and decreased Raf serine-259 phosphorylation. Pharmacological studies revealed that the integrity of lipid raft and the activation of Src, Ras, Raf, ERK, and NF-κB all contributed to JEV-induced TNF-α and IL-1β expression. Pharmacological and biochemical studies further suggested that Src, upon activation, might transmit signals to the Raf/ERK cascades via Ras-dependent and -independent mechanisms that in turn might lead to NF-κB activation. Overall, our results show that the lipid raft might play a role in mediating JEV-initiated Src/Ras/Raf/ERK/NF-κB signaling and TNF-α/IL-1β expression in microglia.
authors.affiliation: Department of Education and Research, Taichung Veterans General Hospital, Taichung, Taiwan. cjchen@vghtc.gov.tw
authors.forename: Chun-Jung
authors.initials: CJ
authors.lastname: Chen
authors.forename: Yen-Chuan
authors.initials: YC
authors.lastname: Ou
authors.forename: Cheng-Yi
authors.initials: CY
authors.lastname: Chang
authors.forename: Hung-Chuan
authors.initials: HC
authors.lastname: Pan
authors.forename: Shih-Yi
authors.initials: SY
authors.lastname: Lin
authors.forename: Su-Lan
authors.initials: SL
authors.lastname: Liao
authors.forename: Shue-Ling
authors.initials: SL
authors.lastname: Raung
authors.forename: Shih-Yun
authors.initials: SY
authors.lastname: Chen
authors.forename: Chen-Jung
authors.initials: CJ
authors.lastname: Chang
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D002478:Cells, Cultured; D009033:Culicidae; D016207:Cytokines / Q000096:biosynthesis*; D004664:Encephalitis Virus, Japanese / Q000302:isolation & purification; D053583:Interleukin-1beta / Q000096:biosynthesis; D021962:Membrane Microdomains / Q000502:physiology; D017628:Microglia / Q000201:enzymology / Q000378:metabolism* / Q000821:virology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis; D019061:src-Family Kinases / Q000378:metabolism* / Q000502:physiology
chemical_list: D016207:Cytokines; D053583:Interleukin-1beta; D014409:Tumor Necrosis Factor-alpha; C490511:Frk protein, rat; D019061:src-Family Kinases