Transcriptomes of control and Chd7-deficient spiral ganglia neurons and cochlear hair cells
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PROVIDER: PRJNA687552 | ENA |
REPOSITORIES: ENA
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Project description:The aim of this study is to identify the gene targets of Chd7 (by RNA-seq) in spiral ganglia neurons and cochlear hair cells. mRNA profiles of FACS-sorted spiral ganglia neurons from 4-day old mice and cochlear hair cells from embryos af 16.5 days in controls, Chd7 heterozygous and Chd7 homozygous mutant animals were generated by paired-end sequencing, in triplicate, using Illumina HiSeq 4000. The sequence reads that passed quality filters were reads were aligned to mouse MM10 genome assembly using HiSAT2 (version 2.1.0) with the default parameters in Galaxy (version 2.1.0). To facilitate quantitative gene expression analysis, aligned reads for each sample were counted using featureCounts (version1.6.4). Differential gene expression analysis was performed using DESeq2 (version 2.11.40.6), applying parametric fit. qRT–PCR validation was performed using SYBR Green and gene specific primers. To identify differentially expressed genes, RPKM values of genes that are not normally expressed in E16.5 hair cells or P4 spiral ganglia neurons were removed from analysis. This resulted in 6910 transcripts for genes expressed in hair cells and 11293 genes expressed in neurons. A pairwise comparison between controls and homozygotes and controls and heterozygotes was performed. Significant thresholds were considered for transcripts with an adjusted p-value (FDR) of ≤0.05 and linear fold change of >2 in either direction. Altered expression of selected genes was confirmed with qRT–PCR as well as the encoded proteins by immunohistochemistry. Analysis of differentially expressed genes uncovered novel targets and potential mechanisms by which Chd7 controls the survival of these cells during normal hearing.
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