DOI: 10.5061/dryad.2bvq83c14
title: Interleukin-17A signaling promotes CD8+ T cell cytotoxicity against West Nile virus infection through enhancing PI3K-mTOR-mediated metabolism

description: West Nile Virus (WNV), a mosquito-borne neurotropic flavivirus, is a major
 cause of viral encephalitis in the United States, posing a continuous
 threat to public health. Unfortunately, no vaccine or specific therapeutic
 intervention is available against WNV infection. Previous studies,
 including ours, demonstrated that interleukin-17A (IL-17A) signaling
 promotes the cytotoxicity of CD8+ T cells to facilitate WNV and parasite
 clearance; however, the molecular mechanism is not understood. IL-17
 receptor C (IL-17RC) is an obligatory co-receptor with IL-17 receptor A
 (IL-17RA) for signaling induced by IL-17A, IL-17A/F, and IL-17F. In this
 study, we found that IL-17RC deficient (Il17rc-/-) mice were more
 susceptible to WNV infection with a higher viral load in the brain than
 wild-type (WT) control mice. The number of infiltrating WNV-specific CD8+
 T cells and the expression levels of cytotoxicity mediators, such as
 perforin, in the T cells in the brain of Il17rc-/- mice were reduced. In
 addition, WNV-specific CD8+ T cells from IL-17RA deficient (Il17ra-/-)
 mice and CD8+ cell-specific Il17ra conditional knockout (cre-KO) mice
 expressed lower levels of perforin than their counterpart controls.
 Moreover, supplementing mouse recombinant IL-17A ex vivo increased the
 perforin production in WNV-specific CD8+ T cells from the WT mice but not
 Il17rc-/- or cre-KO mice. Interestingly, we found that IL-17A signaling
 activated the phosphatidylinositol-3-kinase/mammalian target of rapamycin
 (PI3K-mTOR) signaling pathway in CD8+ T cells, leading to increased
 metabolism of CD8+ T cells to cope with the higher energy demand for WNV
 clearance in the brain. In summary, our findings reveal a novel
 IL-17A-PI3K-mTOR signaling axis in promoting the effector functions of
 CD8+ T cells, suggesting potential broader implications in stimulating
 immune responses to combat WNV and other intracellular infections.
description: RNA sequencing analysis of transcripts of CD8 T cells of IL-17RA
 KO and WT control mice.
description: # Interleukin-17A signaling promotes CD8+ T cell cytotoxicity against West
 Nile virus infection through enhancing PI3K-mTOR-mediated metabolism
 [https://doi.org/10.5061/dryad.2bvq83c14](https://doi.org/10.5061/dryad.2bvq83c14) ## Description of the data and file structure Eight-weeksold *Il17ra-/-*, and WT (C57BL/6J) mice (n = 3) were infected with 100 PFU of WNV via f.p. inoculation, and the spleens were collected on D8 p.i., followed by purification of CD8+ T cells. The transcription profiles of CD8+ T cells were analyzed by RNA sequencing. Library preparation was performed according to the Smart-seq2 protocol (115). Briefly, cDNA was generated from 50ng of total RNA using SuperScript IV reverse transcriptase (Thermo Fisher Scientific, Cat. #18091050) with oligo(dT) priming and template-switching oligo. Double-stranded cDNA was amplified via 12 cycles of PCR using KAPA HiFi HotStart ReadyMix (Roche) and ISPCR oligo. Amplified cDNA was fragmented using the Nextera XT DNA Library Prep Kit (Illumina), with adapter-ligated products size-selected (200–500 bp) via SPRI beads (Beckman Coulter). Then, libraries were quantified by qPCR (KAPA Biosystems) and pooled at equimolar ratios. Paired-end sequencing (2 × 37 bp) was performed on an Illumina NextSeq 500 platform using a High Output Kit v2 (75 cycles). Raw reads were quality-filtered (FastQC v0.11.9) and aligned to the mm10 with TopHat v2.1.1, Bowtie2 v2.2.8, and Samtools v1.3. Counts were normalized to Fragments per kilobase of exon per million mapped fragments (FPKMs), and differential expression analysis was conducted with Cufflinks v2.2.2. ### Files and variables #### File: IL17RA\_RNAseq.xlsx **Description:**  ##### Variables **test_id, gene_id, gene** are the same, are gene symbols, **locus** is genomic coordinates, physical location of the gene in the genome, **`sample_1` & `sample_2`**`,` Names of the compared groups " IL17RA_WT" vs. " IL17RA_KO"), **status**, indicates if the test completed successfully or encountered errors. **`value_1` & `value_2`**, normalized expression values (FPKM, TPM),  **log2(fold_change)**, Log2 ratio of expression values **test_stat**, statistical score, magnitude indicates strength of evidence for differential expression, **p_value**, raw p-value from the statistical test, **q_value**, adjusted p-value for multiple testing **significant**, label indicating significance Code/software ## Access information Other publicly accessible locations of the data: * NA Data was derived from the following sources: * NA