Mettl3/Ythdf2 regulate macrophage inflammation and ROS generation by controlling Pyk2 mRNA stability

Area of Science:

• Immunology
• Molecular Biology
• Epigenetics

Background:

• N6-methyladenosine (m⁶A) is a prevalent RNA modification involved in pathology.
• Mettl3, an m⁶A writer, plays a role in immune response, but its precise mechanisms are unclear.
• Inconsistent findings highlight the need to identify Mettl3's specific targets in inflammation.

Purpose of the Study:

• To investigate the role of Mettl3 in cellular inflammatory response.
• To identify Mettl3 target genes involved in inflammation.
• To elucidate the molecular mechanism by which Mettl3 regulates inflammatory pathways.

Main Methods:

• RNA sequencing (RNA-seq) in Mettl3-ablated macrophages.
• m⁶A mapping and RIP-PCR.
• Analysis of inflammatory markers, ROS generation, and signaling pathway activation (MAPKs, AKT).
• Validation using a Pyk2 inhibitor.

Main Results:

• Mettl3 ablation enhanced LPS-induced IL-6, TNF-α, NOXs expression, ROS generation, and MAPK/AKT phosphorylation.
• Pyk2 was identified as a potential Mettl3 target gene.
• Mettl3 and Ythdf2 depletion increased Pyk2 expression and mRNA stability.
• Ythdf2 directly targets Pyk2 in a Mettl3-dependent manner.
• Pyk2 inhibition reversed Mettl3 knockdown-induced inflammatory responses.

Conclusions:

• Mettl3 regulates Pyk2 mRNA stability and expression through Ythdf2.
• This regulation impacts MAPKs and AKT signaling, NOXs upregulation, and subsequent pro-inflammatory cytokine and ROS production.
• Mettl3 acts as a negative regulator of inflammatory responses by controlling Pyk2.