Dataset Information

Mettl14-Mediated m⁶A Modification Facilitates Liver Regeneration by Maintaining Endoplasmic Reticulum Homeostasis.

ABSTRACT:

Background & aims

N⁶-methyladenosine (m⁶A), the most prevalent and dynamic posttranscriptional methylation modification of mammalian mRNA, is involved in various biological processes, but its role in liver regeneration has not been characterized.

Methods

We first conducted transcriptome-wide m⁶A mRNA sequencing and characterized the expression pattern of m⁶A in regenerating mouse liver. Next, we generated hepatocyte-specific Mettl3- or Mettl14-deficient mice and investigated their role in liver regeneration. A series of biochemical experiments in vitro and in vivo was further performed to investigate potential mechanisms.

Results

We identified an overwhelming proportion of m⁶A-modified genes with initially up-regulated and subsequently down-regulated m⁶A levels as liver regeneration progressed. Loss of Mettl14 but not of Mettl3 resulted in markedly disrupted liver regeneration, and Mettl14-ablated hepatocytes were arrested in the G1 phase of the cell cycle. Most strikingly, the Mettl14-ablated regenerating liver exhibited extensive parenchymal necrosis. mRNA transcripts, such as Hsp90b1, Erp29, Stt3a, P4hb, and Lman1, encoding proteins involved in polypeptide processing and the endoplasmic reticulum (ER) stress response, were m⁶A-hypomethylated, and their mRNA and protein levels were subsequently decreased, resulting in unresolved ER stress, hepatocyte death, and inhibited proliferation.

Conclusions

We demonstrate the essential role of Mettl14 in facilitating liver regeneration by modulating polypeptide-processing proteins in the ER in an m⁶A-dependent manner.

SUBMITTER: Cao X

PROVIDER: S-EPMC8261664 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND & AIMS:Disruption to endoplasmic reticulum (ER) calcium homeostasis has been implicated in obesity, however, the ability to longitudinally monitor ER calcium fluctuations has been challenging with prior methodologies. We recently described the development of a Gaussia luciferase (GLuc)-based reporter protein responsive to ER calcium depletion (GLuc-SERCaMP) and investigated the effect of a high fat diet on ER calcium homeostasis. METHODS:A GLuc-based reporter cell line was treated with palmitate, a free fatty acid. Rats intrahepatically injected with GLuc-SERCaMP reporter were fed a cafeteria diet or high fat diet. The liver and plasma were examined for established markers of steatosis and compared to plasma levels of SERCaMP activity. RESULTS:Palmitate induced GLuc-SERCaMP release in vitro, indicating ER calcium depletion. Consumption of a cafeteria diet or high fat pellets correlated with alterations to hepatic ER calcium homeostasis in rats, shown by increased GLuc-SERCaMP release. Access to ad lib high fat pellets also led to a corresponding decrease in microsomal calcium ATPase activity and an increase in markers of hepatic steatosis. In addition to GLuc-SERCaMP, we have also identified endogenous proteins (endogenous SERCaMPs) with a similar response to ER calcium depletion. We demonstrated the release of an endogenous SERCaMP, thought to be a liver esterase, during access to a high fat diet. Attenuation of both GLuc-SERCaMP and endogenous SERCaMP was observed during dantrolene administration. CONCLUSIONS:Here we describe the use of a reporter for in vitro and in vivo models of high fat diet. Our results support the theory that dietary fat intake correlates with a decrease in ER calcium levels in the liver and suggest a high fat diet alters the ER proteome. Lay summary: ER calcium dysregulation was observed in rats fed a cafeteria diet or high fat pellets, with fluctuations in sensor release correlating with fat intake. Attenuation of sensor release, as well as food intake was observed during administration of dantrolene, a drug that stabilizes ER calcium. The study describes a novel technique for liver research and provides insight into cellular processes that may contribute to the pathogenesis of obesity and fatty liver disease.

Dataset Information

Mettl14-Mediated m⁶A Modification Facilitates Liver Regeneration by Maintaining Endoplasmic Reticulum Homeostasis.

Background & aims

Methods

Results

Conclusions

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OmicsDI is part of the ELIXIR infrastructure

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OmicsDI is part of the ELIXIR infrastructure

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Mettl14-Mediated m⁶A Modification Facilitates Liver Regeneration by Maintaining Endoplasmic Reticulum Homeostasis.