The crossroads of RNA methylation and ferroptosis: Implications for disease pathogenesis

Abstract

Post-transcriptional regulation constitutes an important mechanism for governing gene expression across diverse biological processes.¹ It is now appreciated that multiple types of methylation in mRNA provide post-transcriptional regulation to control mRNA fates. N⁶-methyladenosine (m⁶A) methylation is one of the most prevalent internal RNA modifications.² m⁶A methylation is involved in various aspects of post-transcriptional regulation, such as the control of mRNA decay, translation and splicing. Accumulating evidence has shown that m⁶A methylation controls numerous biological processes, such as cell development, immune response and metabolic control.^{3, 4}

The intricate interplay between RNA m⁶A methylation and programmed cell death or apoptosis, represents an expanding area of research in molecular biology. Studies have unveiled connections between m⁶A methylation and the regulation of apoptotic pathways. Specifically, m⁶A modifications have been found to modulate the expression of key genes involved in apoptosis, thereby exerting a regulatory influence on cell survival and death decisions. This regulatory role is particularly evident in the context of cancer biology, where dysregulation of m⁶A methylation can lead to aberrant apoptotic signaling, contributing to tumour progression.⁵ Studying the precise mechanisms by which RNA methylation influences apoptotic pathways has the potential to reveal new therapeutic strategies for diseases with disrupted cell death regulation.

A new form of programmed cell death called ferroptosis adds a captivating dimension to the complex landscape of cell death regulation.⁶ Unlike classical apoptosis, ferroptosis is a distinct form of regulated cell death, characterised by the iron-dependent accumulation of lipid peroxides, ultimately leading to membrane damage and cell death. Despite the growing recognition of ferroptosis as a significant cellular process with implications in various pathological conditions, such as neurodegenerative diseases and cancer, our understanding of the interplay between m⁶A methylation and ferroptosis remains in its infancy.

In a recent study by Zhang et al.,⁷ a novel connection was established between RNA m⁶A modification, ferroptosis and the development of acute lung injury (ALI), a sudden and severe lung condition characterised by inflammation and increased lung tissue permeability.⁸ ALI often results in impaired oxygen exchange and hypoxaemia, necessitating prompt medical intervention to prevent progression to acute respiratory distress syndrome (ARDS). Initially, the research explored the role of neutrophil extracellular traps (NETs) in the pathogenesis of sepsis-associated ALI (SI-ALI). The study revealed a significant correlation between the severity of the disease and the formation of NETs in the lungs of hospitalised patients. Subsequently, the study employed an in vivo SI-ALI model utilising the cecal ligation and puncture mouse model, which induced the recruitment of neutrophils and NETs formation, both of which appeared to play a significant role in the pathogenesis. Interestingly, the study detected an increase in the concentration of MDA (malondialdehyde), an end product of lipid peroxidation, as well as elevated levels of reactive oxygen species (ROS) and ferrous iron, along with reduced GPX4 and glutathione (GSH) levels—all indicative of ferroptosis. Notably, these changes in ferroptosis markers showed partial dependence on neutrophils and NETs formation. In vitro experiments showed that purified NETs had a similar effect on human alveolar epithelial cells (HPAEpiCs), and the effects of NETs were cancelled by ferrostatin-1, an inhibitor of ferroptosis. These findings suggest that NET-induced ferroptosis is a key driver of SI-ALI.

Next, Zhang et al. investigated the contribution of m⁶A methylation to the control of ferroptosis and the development of SI-ALI, as the authors previously reported that the treatment of alveolar epithelial cells with NETs enhanced m⁶A methylation.⁹ Their RNA-seq analysis revealed an upregulation in the expression of the pivotal m⁶A methyltransferase METTL3 and the m⁶A binding protein IGF2BP2¹⁰ in response to NETs treatment. The increased expression of METTL3 is likely dependent on p300-H3K27ac axis. The ablation of METTL3 conferred resistance to ferroptosis induced by NETs treatment in vitro, underscoring its pivotal role in this cell death process. Specifically, METTL3 knockdown in vitro resulted in increased expression of GPX4, a key suppressor of ferroptosis. Furthermore, alveolar epithelial cell-specific depletion of METTL3 in vivo was protected from SI-ALI. These findings suggest that METTL3-dependent m⁶A regulation contributes to ferroptosis induction and the development of NETs-induced SI-ALI.

The study further delved into the molecular mechanisms underlying the regulation of ferroptosis by METTL3. By conducting an integrative analysis that combined RNA-seq with methylated RNA immunoprecipitation sequencing (MeRIP-seq), the researchers identified hypoxia-inducible factor (HIF)-1α as a crucial target for methylation by METTL3 in the control of ferroptosis. METTL3 was found to induce the expression of HIF-1α in a manner dependent on m⁶A and IGF2BP2. Mechanistically, the deposition of m⁶A on HIF-1α mRNA facilitated the recruitment of IGF2BP2, thereby enhancing the stability of this specific mRNA. HIF-1α was observed to downregulate the expression of GPX4, implying that the accumulation of HIF-1α acts as an upstream regulator of ferroptosis induction in this particular context.

HIF-1α serves as a pivotal transcription factor responsible for initiating responses to hypoxia and/or inflammation by stimulating the expression of glycolytic enzymes and angiogenic factors.¹¹ In agreement with the observed shifts in HIF-1α expression, METTL3 knockdown led to the downregulation of several glycolytic enzymes and an abrogation of glycolytic activity. Although the precise mechanisms through which the metabolic alterations induced by HIF-1α influence ferroptosis remain largely elusive, these findings underscore the emergence of a novel interplay between mRNA methylation, ferroptosis and the pathogenesis of ALI.

The recent discovery by Zhang et al. prompts a series of intriguing questions: Does METTL3 control ferroptosis in different contexts, including cancers and cardiovascular and neurodegenerative diseases? How is NETs treatment sensed by alveolar epithelial cells, leading to the induction of METTL3 expression? What is the biological significance of linking RNA methylation as an upstream regulator of ferroptosis induction in a broader context? This study serves as an inspiring catalyst for further research, which promises to shed more light on the molecular link between these two processes and elucidate their broader implications in various physiological and pathological settings.

The author declares no conflict of interest.