Sphingosine kinase 2 is a chikungunya virus host factor co-localized with the viral replication complex

Abstract

Chikungunya virus (CHIKV) is a re-emerging alphavirus which causes severe and prolonged arthralgic febrile illness. The recent global spread of the virus and lack of approved therapeutic options makes it imperative to gain greater insight into the molecular mechanisms underlying CHIKV pathogenesis, in particular host factors recruited by the virus. In the current study, we identify sphingosine kinase 2 (SK2) as a CHIKV host factor co-localized with the viral replication complex (VRC) during infection. SK2 was demonstrated to co-localize with viral RNA and nonstructural proteins. Targeted impairment of SK2 expression or function significantly inhibited CHIKV infection. Furthermore, affinity purification-mass spectrometry studies revealed that SK2 associates with a number of proteins involved in cellular gene expression specifically during viral infection, suggesting a role in replication. Collectively these results identify SK2 as a novel CHIKV host factor.

Validation of commercially available sphingosine kinase 2 antibodies for use in immunoblotting, immunoprecipitation and immunofluorescence

“…We have examined a rabbit polyclonal SK2 antibody from Proteintech, which is raised against amino acids 266–618 of recombinant human SK2a, and a rabbit polyclonal SK2 antibody from ECM Biosciences, which is raised against a synthetic peptide corresponding to amino acids 36–52 of human SK2a. The Proteintech SK2 antibody has been previously utilized in one publication for IB 11 , and the ECM Biosciences SK2 antibody has been used in multiple publications for IB 12– 15 and for IF 16 .…”

In silico study on anti-Chikungunya virus activity of hesperetin

“…NsP3-mediated SPK2 re-localisation to a puncta in the cytoplasm following an infection suggested the involvement of SPK2 in CHIKV replication. Close association of SPK2 with host proteins for mRNA processing and gene expression further link SPK2 with CHIKV replication process (Reid et al, 2015). Our findings suggest that inhibitory effect of hesperetin could be contributed by both direct activity on the virus proteins as well as cellular factors vital for the virus replication.…”

“…Previous studies have shown close association between SPK/S1P pathway and important cellular processes such as cell growth, inflammation and immune responses, with both types of SPKs having the opposing effects (Maceyka et al, 2005; Okada et al, 2005; Olivera et al, 1999; Snider, Alexa Orr Gandy & Obeid, 2010). Interestingly, in addition to its contribution towards the development of degenerative diseases in humans, SPKs also play important roles in several viral infections, including Dengue and Influenza A viruses for SPK1 as well as Hepatitis C virus, Kaposi’s sarcoma-associated herpes virus, and most importantly for our study, CHIKV for SPK2 (Clarke et al, 2016; Couttas et al, 2014; Dai et al, 2014; Reid et al, 2015; Seo et al, 2013; Yamane et al, 2014). …”

Transforming Sphingosine Kinase 1 Inhibitors into Dual and Sphingosine Kinase 2 Selective Inhibitors: Design, Synthesis, and in Vivo Activity

“…12 S1P interacts with proteins both within and outside of the cell. As an extracellular ligand, S1P promotes cell migration and survival by binding to five G-protein coupled receptors, S1P 1-5 .…”

“…SphK1 is a cytosolic enzyme that promotes cell survival and proliferation, 21, 22 whereas some SphK2 is found in the nucleus 17, 18 but can relocate to the cytosol on phosphorylation. 23, 24 Depending on this localization, SphK2 can promote either apoptosis 12,13 or cell proliferation. 17, 18, 25, 26 Although isotype-specific SphK null mice are viable, fertile, and phenotypically unremarkable, 27 SphK1-null mice have about a two-fold reduction in blood S1P levels while SphK2-null mice have 2-4 fold increased blood S1P levels.…”