The MRN complex maintains the biliary-derived hepatocytes in liver regeneration through ATR-Chk1 pathway.

When the proliferation of residual hepatocytes is prohibited, biliary epithelial cells (BECs) transdifferentiate into nascent hepatocytes to accomplish liver regeneration. Despite significant interest in transdifferentiation, little is known about the maintenance of nascent hepatocytes in post-injured environments. Here, we perform an N-ethyl-N-nitrosourea (ENU) forward genetic screen and identify a mutant containing a nonsense mutation in the gene nibrin (nbn), which encodes a component of the Mre11-Rad50-Nbn (MRN) complex that activates DNA damage response (DDR). The regenerated hepatocytes cannot be maintained and exhibit apoptosis in the mutant. Mechanistically, the nbn mutation results in the abrogation of ATR-Chk1 signaling and accumulations of DNA damage in nascent hepatocytes, which eventually induces p53-mediated apoptosis. Furthermore, loss of rad50 or mre11a shows similar phenotypes. This study reveals that the activation of DDR by the MRN complex is essential for the survival of BEC-derived hepatocytes, addressing how to maintain nascent hepatocytes in the post-injured environments.

Rngtt governs biliary-derived liver regeneration initiation by transcriptional regulation of mTORC1 and Dnmt1 in zebrafish.

In cases of end-stage liver diseases, the proliferation of existing hepatocytes is compromised, a feature of human chronic liver disease, in which most hepatocytes are dysfunctional. So far, liver transplantation represents the only curative therapeutic solution for advanced liver diseases, and the shortage of donor organs leads to high morbidity and mortality worldwide. The promising treatment is to prompt the biliary epithelial cells (BECs) transdifferentiation. However, the critical factors governing the initiation of BEC-derived liver regeneration are largely unknown. The zebrafish has advantages in large-scale genetic screens to identify the critical factors involved in liver regeneration. Here, we combined N-ethyl-N-nitrosourea screen, positional cloning, transgenic lines, antibody staining, and in situ hybridization methods and identified a liver regeneration defect mutant ( lrd ) using the zebrafish extensive liver injury model. Through positional cloning and genomic sequencing, we mapped the mutation site to rngtt . Loss of rngtt leads to the defects of BEC dedifferentiation, bipotential progenitor cell activation, and cell proliferation in the initiation stage of liver regeneration. The transdifferentiation from BECs to hepatocytes did not occur even at the late stage of liver regeneration. Mechanically, Rngtt transcriptionally regulates the attachment of mRNA cap to mTOR complex 1 (mTORC1) components and dnmt1 to maintain the activation of mTORC1 and DNA methylation in BECs after severe liver injury and prompt BEC to hepatocyte conversion. Furthermore, rptor and dnmt1 mutants displayed the same liver regeneration defects as rngtt mutation. In conclusion, our results suggest Rngtt is a new factor that initiates BEC-derived liver regeneration.

DNA methylation maintenance at the p53 locus initiates biliary-mediated liver regeneration.

In cases of extensive liver injury, biliary epithelial cells (BECs) dedifferentiate into bipotential progenitor cells (BPPCs), then redifferentiate into hepatocytes and BECs to accomplish liver regeneration. Whether epigenetic regulations, particularly DNA methylation maintenance enzymes, play a role in this biliary-mediated liver regeneration remains unknown. Here we show that in response to extensive hepatocyte damages, expression of dnmt1 is upregulated in BECs to methylate DNA at the p53 locus, which represses p53 transcription, and in turn, derepresses mTORC1 signaling to activate BEC dedifferentiation. After BEC dedifferentiation and BPPC formation, DNA methylation at the p53 locus maintains in BPPCs to continue blocking p53 transcription, which derepresses Bmp signaling to induce BPPC redifferentiation. Thus, this study reveals promotive roles and mechanisms of DNA methylation at the p53 locus in both dedifferentiation and redifferentiation stages of biliary-mediated liver regeneration, implicating DNA methylation and p53 as potential targets to stimulate regeneration after extensive liver injury.