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Extended JAK activation and delayed STAT1 dephosphorylation contribute to the distinct signaling profile of CNS neurons exposed to interferon-gamma.

ABSTRACT: Although interferon-gamma (IFN-?) plays a critical role in the noncytolytic elimination of many neurotropic viral infections, the signaling response to this cytokine has not been extensively characterized in primary CNS neurons. We previously demonstrated that the IFN-? response at the signaling and gene expression levels is temporally extended in primary mouse hippocampal neurons, as compared to the transient response of primary mouse embryonic fibroblasts (MEF). We hypothesize that the protracted kinetics of STAT1 phosphorylation in IFN-?-treated neurons are due to extended receptor activation and/or delayed STAT1 dephosphorylation in the nucleus. Here, we show that in response to IFN-?, the Janus kinases (JAK1/JAK2) associated with the neuronal IFN-? receptor complex remain active for an extended period as compared to MEF. Experimental inactivation of JAK1/JAK2 in neurons after IFN-? treatment did not reverse the extended STAT1 phosphorylation phenotype. These results suggest that the extended kinetics of neuronal IFN-? signaling are a product of distinct negative feedback mechanisms operating at both the receptor and within the nucleus.

SUBMITTER: Podolsky MA

PROVIDER: S-EPMC3678282 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature

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Extended JAK activation and delayed STAT1 dephosphorylation contribute to the distinct signaling profile of CNS neurons exposed to interferon-gamma.

Podolsky Michael A MA Solomos Andreas C AC Durso Lisa C LC Evans Stephanie M SM Rall Glenn F GF Rose R Wesley RW

Journal of neuroimmunology 20120704 1-2

Although interferon-gamma (IFN-γ) plays a critical role in the noncytolytic elimination of many neurotropic viral infections, the signaling response to this cytokine has not been extensively characterized in primary CNS neurons. We previously demonstrated that the IFN-γ response at the signaling and gene expression levels is temporally extended in primary mouse hippocampal neurons, as compared to the transient response of primary mouse embryonic fibroblasts (MEF). We hypothesize that the protrac ...[more]

PMID: 22769061

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Project description:Signal transducer and activator of transcription (STAT1)1 gain of function (GOF) pathogenic variants have been associated with increased levels of phosphorylated STAT1 and STAT1-dependent cellular responses. Delayed dephosphorylation was proposed as the underlying mechanism leading to the characteristically raised pSTAT1 levels. We examined the levels of STAT1 protein and message as well as rates of STAT1 phosphorylation, dephosphorylation, and degradation associated with STAT1 GOF pathogenic variants. Fresh peripheral blood mononuclear cells (PBMC) from 14 STAT1 GOF patients carrying 10 different pathogenic variants in the coiled-coil, DNA binding, and SH2 domains and healthy donors were used to study STAT1 levels and phosphorylation (pSTAT1) following IFNγ and IFNα stimulation. STAT1 protein levels were measured by flow cytometry and immunoblot. STAT1 mRNA levels were measured using quantitative reverse transcription PCR. STAT1 protein degradation was studied using cycloheximide. Patient IFNγ and IFNα induced peak pSTAT1 was higher than in healthy controls. The velocity of pSTAT1 dephosphorylation after treatment of IFNγ stimulated CD14+ monocytes with the Janus Kinase (JAK)-inhibitor ruxolitinib was significantly faster in patient cells. STAT1 protein levels in patient CD14+ monocytes and CD3+ T cells were higher than in healthy donors. There was a strong and positive correlation between CD14+ STAT1 protein levels and peak pSTAT1 levels. Patient fresh PBMC STAT1 mRNA levels were increased at rest and after 16 h of incubation. STAT1 protein degradation was similar in patient and healthy volunteer cells. Patient IFNγ receptors 1 and 2 and JAK2 levels were normal. One patient in our cohort was treated with the oral JAK inhibitor ruxolitinib. Treatment was associated with normalization of both STAT1 protein and peak pSTAT1 levels. After JAK inhibitor treatment was stopped the patient's CD14+ monocyte STAT1 protein and peak phosphorylation levels increased proportionally. These findings suggest that patients with STAT1 GOF mutations have higher levels of total STAT1 protein, leading to high levels of pSTAT1 after stimulation, despite rapid STAT1 dephosphorylation and normal degradation.

Dataset Information

Extended JAK activation and delayed STAT1 dephosphorylation contribute to the distinct signaling profile of CNS neurons exposed to interferon-gamma.

Publications

Extended JAK activation and delayed STAT1 dephosphorylation contribute to the distinct signaling profile of CNS neurons exposed to interferon-gamma.

Similar Datasets

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