Dataset Information

Sox9 Is a Modifier of the Liver Disease Severity in a Mouse Model of Alagille Syndrome.

ABSTRACT:

Background and aims

Alagille syndrome (ALGS) is a multisystem developmental disorder characterized by bile duct (BD) paucity, caused primarily by haploinsufficiency of the Notch ligand jagged1. The course of the liver disease is highly variable in ALGS. However, the genetic basis for ALGS phenotypic variability is unknown. Previous studies have reported decreased expression of the transcription factor SOX9 (sex determining region Y-box 9) in late embryonic and neonatal livers of Jag1-deficient mice. Here, we investigated the effects of altering the Sox9 gene dosage on the severity of liver disease in an ALGS mouse model.

Approach and results

Conditional removal of one copy of Sox9 in Jag1^+/- livers impairs the biliary commitment of cholangiocytes and enhances the inflammatory reaction and liver fibrosis. Loss of both copies of Sox9 in Jag1^+/- livers further worsens the phenotypes and results in partial lethality. Ink injection experiments reveal impaired biliary tree formation in the periphery of P30 Jag1^+/- livers, which is improved by 5 months of age. Sox9 heterozygosity worsens the P30 biliary tree phenotype and impairs the partial recovery in 5-month-old animals. Notably, Sox9 overexpression improves BD paucity and liver phenotypes in Jag1^+/- mice without ectopic hepatocyte-to-cholangiocyte transdifferentiation or long-term liver abnormalities. Notch2 expression in the liver is increased following Sox9 overexpression, and SOX9 binds the Notch2 regulatory region in the liver. Histological analysis shows a correlation between the level and pattern of SOX9 expression in the liver and outcome of the liver disease in patients with ALGS.

Conclusions

Our results establish Sox9 as a dosage-sensitive modifier of Jag1^+/- liver phenotypes with a permissive role in biliary development. Our data further suggest that liver-specific increase in SOX9 levels is a potential therapeutic approach for BD paucity in ALGS.

SUBMITTER: Adams JM

PROVIDER: S-EPMC7048647 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background & aimsAlagille syndrome is a genetic disorder characterized by cholestasis, ocular abnormalities, characteristic facial features, heart defects, and vertebral malformations. Most cases are associated with mutations in JAGGED1 (JAG1), which encodes a Notch ligand, although it is not clear how these contribute to disease development. We aimed to develop a mouse model of Alagille syndrome to elucidate these mechanisms.MethodsMice with a missense mutation (H268Q) in Jag1 (Jag1+/Ndr mice) were outbred to a C3H/C57bl6 background to generate a mouse model for Alagille syndrome (Jag1Ndr/Ndr mice). Liver tissues were collected at different timepoints during development, analyzed by histology, and liver organoids were cultured and analyzed. We performed transcriptome analysis of Jag1Ndr/Ndr livers and livers from patients with Alagille syndrome, cross-referenced to the Human Protein Atlas, to identify commonly dysregulated pathways and biliary markers. We used species-specific transcriptome separation and ligand-receptor interaction assays to measure Notch signaling and the ability of JAG1Ndr to bind or activate Notch receptors. We studied signaling of JAG1 and JAG1Ndr via NOTCH 1, NOTCH2, and NOTCH3 and resulting gene expression patterns in parental and NOTCH1-expressing C2C12 cell lines.ResultsJag1Ndr/Ndr mice had many features of Alagille syndrome, including eye, heart, and liver defects. Bile duct differentiation, morphogenesis, and function were dysregulated in newborn Jag1Ndr/Ndr mice, with aberrations in cholangiocyte polarity, but these defects improved in adult mice. Jag1Ndr/Ndr liver organoids collapsed in culture, indicating structural instability. Whole-transcriptome sequence analyses of liver tissues from mice and patients with Alagille syndrome identified dysregulated genes encoding proteins enriched at the apical side of cholangiocytes, including CFTR and SLC5A1, as well as reduced expression of IGF1. Exposure of Notch-expressing cells to JAG1Ndr, compared with JAG1, led to hypomorphic Notch signaling, based on transcriptome analysis. JAG1-expressing cells, but not JAG1Ndr-expressing cells, bound soluble Notch1 extracellular domain, quantified by flow cytometry. However, JAG1 and JAG1Ndr cells each bound NOTCH2, and signaling from NOTCH2 signaling was reduced but not completely inhibited, in response to JAG1Ndr compared with JAG1.ConclusionsIn mice, expression of a missense mutant of Jag1 (Jag1Ndr) disrupts bile duct development and recapitulates Alagille syndrome phenotypes in heart, eye, and craniofacial dysmorphology. JAG1Ndr does not bind NOTCH1, but binds NOTCH2, and elicits hypomorphic signaling. This mouse model can be used to study other features of Alagille syndrome and organ development.