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Dynamic gene expression patterns during in vivo maturation of mouse hepatoblasts Lee, Sam Chi-Hang


Hepatoblasts are bipotential fetal liver cells that differentiate into hepatocytes and cholangiocytes, the parenchymal cell types in the adult liver. While genes that regulate hepatoblast differentiation have been identified, less is known about the underlying regulatory networks. Transcriptome studies using mouse whole fetal livers have been undertaken to decipher those networks. However, as 70-80% of cells in the fetal liver are hematopoietic, the resulting transcriptome data were biased toward genes expressed in blood cells and not maturing hepatoblasts. To address this, I performed a temporal transcriptome analysis using highly purified mouse fetal hepatoblasts. First, I showed that the cell surface molecule DLK1 is specifically expressed on hepatoblasts throughout fetal development. Then, fetal liver DLK1⁺ cells were isolated on embryonic days 10.5, 12.5, 14.5 and 16.5, and their respective transcriptomes were captured in Tag-seq libraries. The Tag-seq data were specifically enriched for hepatic genes, and liver-enriched transcription factors that were detected at marginal levels in previous fetal liver microarray studies were now readily observed. Furthermore, while genes associated with hepatoblast identity were consistently observed in all four DLK1⁺ libraries, cholangiocyte genes showed a gradual decrease over time, implicating DLK1 to be repressed during cholangiocyte differentiation. To identify the temporal gene expression profiles underlying hepatoblast maturation, 5449 highly expressed genes in the Tag-seq libraries were subjected to K-means clustering to generate 14 gene groups with distinct temporal expression patterns. Gene Ontology terms were differentially enriched in the gene clusters, suggesting temporally co-regulated genes to share biological functions. Further analysis was performed on HMGA2, a transcription factor with high transcript expression in early hepatoblasts. Expression of the HMGA2 protein was prominent in nascent hepatoblasts but became restricted to a small subset of hepatoblasts over time. Hmga2 mutant embryos showed a slight reduction of hepatoblasts in the fetal liver, and high Hmga2 was correlated with suppressed expression of the hepatic transcription factor HNF4A in liver cells lines. Overall, this study provides the first genome-wide, temporal analysis of the mouse hepatoblast transcriptome, and the Tag-seq libraries provide a strong foundation for understanding genetic programs that drive in vivo hepatoblast differentiation.

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