RNA Sequencing Allows for Differential Gene Expression Analysis of Infrequently Dividing and Frequently Dividing (IDC/FDC) Oral Epithelial Stem Cells

ABSTRACT: Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare palatal epithelial basal cell transcriptomes between infrequently and frequently dividing cells (IDC/FDCs). Methods: RNA was isolated from 200-1000 palatal basal epithelial cells and libraries prepared using the SoLo RNA-Seq library preparation kit (NuGEN). mRNA profiles of 14-day doxycycline chased K5-tTA;tetO-H2B-GFP (containing populations of slow-cycling GFP-HI and fast-cycling GFP-LO) mice were generated with 6 matched samples using an Illumina HiSeq4000. RNA Sequencing Allows for Differential Gene Expression Analysis of infrequently dividing and frequently dividing cells (IDC/FDC). RNA-seq fastq files were trimmed and aligned to the latest mouse genome, mm10 (GRCm38/GCA_000001635.2) using BBMap. A matrix of the data was generated using featureCounts. RNAseq data was analyzed with DESeq2. All statistical analyses were performed using R version 3.4.1 and RStudio version 1.0.143. Validation was performed using lineage tracing and immunofluorescence immunohistochemistry. Results: Using an optimized data analysis workflow, approximately 2% of the transcripts showed differential expression between the GFP-HI and GFO-LO palatal basal epithelial cell with a p value <0.05. Analysis of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to oral epithelial stem cell, such as Lrig1 and Igfbp5 as followed up on in the associated manuscript. Conclusions: Our study represents the first detailed analysis of palatal basal epithelial stem cell transcriptomes, with biologic replicates, generated by RNA-seq technology. Our results show GFP-LO infrequently dividing cells are express a distinct transcriptional profile compared to frequently dividing neighbor cells such as Lrig1 and Igfbp5.

ORGANISM(S): Mus musculus

PROVIDER: GSE139146 | GEO | 2019/10/22

REPOSITORIES: GEO