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Next Generation Sequencing Facilitates Quantitative Analysis of Runx2fl/fl and Gli1-CreERT2;Runx2fl/fl Incisor Transcriptomes

ABSTRACT: Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived incisor transcriptome profiling (RNA-seq) between Runx2fl/fl and Gli1-CreERT2;Runx2fl/fl mice to identify the potential target of Runx2. Methods: Incisor mRNA profiles from one week after induction of one-month-old Runx2fl/fl and Gli1-CreERT2;Runx2fl/fl mice were generated by deep sequencing, in triplicate, using Illumina NextSeq500. Results: Using an optimized data analysis workflow, we mapped about 68 million sequence reads per sample to the mouse genome ( mm10) and identified 80,214 transcripts in th incisors of Runx2fl/fl and Gli1-CreERT2;Runx2fl/fl mice with Partek E/M workflow. Five hundred and eleven differentially regulated genes were identified (＞2-fold, p＜0.05), of which 299 were upregulated and 212 were downregulated.

ORGANISM(S): Mus musculus

PROVIDER: GSE146332 | GEO | 2020/03/05

REPOSITORIES: GEO

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Project description:The tooth is a convenient experimental model for the study of the basic mechanisms of organ development, including differentiation, cellular interaction, morphogenesis, and production and mineralization of extracellular matrices. The rodent incisor teeth grow continuously and exhibit all stages of tooth development at any time. This characteristic makes them convenient models for the study of enamel formation, which occurs in several distinct stages along the tooth axis. The distribution and structure of mouse incisor enamel resemble that of the rat. The enamel covers only the labial aspect of the tooth, and can be divided into four layers: a thin inner prism-free layer, inner enamel with prism decussation, i.e. transverse rows of prisms with prisms inclined medially and laterally in alternate rows, outer enamel with parallel prisms inclined incisally and a thin superficial prism-free layer. Recently, we have described how this elaborate organization of the enamel is established in the initial enamel formed on the unerupted and unworn incisal tip of the incisors. The very first enamel formed, i.e. the most incisally situated enamel, is always prism-free, corresponding to the superficial layer in fully established enamel, although thicker. Going in apical direction, i.e. in the direction of initiation of new enamel formation, isolated prisms appear among the crystals continuous with the prism-free zone crystals. These initial prisms are in general inclined incisally, corresponding to the prisms in the outer enamel layer in the fully established enamel. Inner enamel with the characteristic decussation pattern is added somewhat further apically and increases in thickness with increasing enamel thickness. Based on the structure of the enamel that they produce, the ameloblasts producing the initial, thin, prism-free enamel at the incisal tip of the unworn mouse incisor are probably differently configured (e.g. lacking Tomes' processes), probably differently organized (e.g. not organized in transverse rows moving sideways in opposite directions), and probably have a shorter life-span than the more apically situated ameloblasts which produce thicker enamel with the full four-layered configuration of mouse incisor enamel. For this reason it would be of interest to compare the gene expression profile of the incisal tip segment where prism-free enamel is being formed with the profile of the immediately adjacent segment where enamel with all four layers is being formed. As long as the incisor is unworn enamel formation and gene expression in these segments will likely reflect mechanistic differences between these segments as regards enamel biosynthesis. MicroRNAs (miRNA) are a class of non-coding RNAs that regulate gene expression at a post-transcriptional level, and are considered as important regulatory molecules during foetal development. By binding to target mRNA, miRNA induce mRNA decay or translation repression. Recent bioinformatic predictions of miRNA targets in vertebrates indicate that hundreds of miRNAs are responsible for regulating the expression of up to 30% of the human protein-coding genes, and miRNAs have recently also been shown to regulate expression of hundreds of mRNAs in cultured cells. Recently, miRNA expression profiles of the developing murine molar tooth germ and submandibular salivary gland have been established. We here report expression profiling of miRNAs present in the two adjacent segments of the incisor tooth germ on which we also have carried out mRNA expression profiling. The results suggest that also miRNAs are abundantly, and differentially, expressed during development of the incisor tooth germ in mouse. By analogy with the mRNA data also miRNA expression is dynamic, with respect to both the two incisor segments investigated and to the developmental stage of the incisor.

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Next Generation Sequencing Facilitates Quantitative Analysis of Runx2fl/fl and Gli1-CreERT2;Runx2fl/fl Incisor Transcriptomes

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