Alternative mRNA Polyadenylation Regulates Macrophage Hyperactivation via the Autophagy Pathway
NUDT21 Regulates Macrophage Hyperactivation via the Autophagy Pathway
Academic Background
The inflammatory response is a critical component of the body’s immune defense, facilitating pathogen clearance and tissue repair. However, dysregulation of the inflammatory response can lead to chronic inflammatory diseases such as inflammatory bowel disease (IBD), rheumatoid arthritis (RA), psoriasis, and systemic lupus erythematosus (SLE). These diseases are often accompanied by aberrant immune system activation and chronic inflammation, which, in severe cases, may progress to life-threatening conditions such as hemophagocytic lymphohistiocytosis (HLH). Excessive macrophage activation is a hallmark of these inflammatory conditions, and their dysregulated function can lead to tissue damage, sustained inflammation, and disease exacerbation. Hence, studying the complex regulatory mechanisms governing macrophage activation is essential for implementing timely measures to prevent excessive inflammatory responses.
On a molecular level, precise transcriptional regulation and alterations in RNA metabolism are tightly connected to macrophage activation states and cytokine production. RNA metabolism includes RNA synthesis, processing, and degradation, among other processes. Alternative polyadenylation (APA) is a key process in RNA metabolism that generates mRNA isoforms with varying 3’ untranslated region (3’UTR) lengths by selecting different polyadenylation sites (PAS). APA is primarily categorized into two types: 3’UTR-APA, which adjusts 3’UTR length without altering coding regions, and intronic-APA, which may produce truncated proteins. Research has demonstrated that 3’UTR shortening occurs during macrophage differentiation or under viral stimulation. However, the specific roles of APA regulators in macrophage activation remain unclear.
Paper Overview
This study was conducted by Yunzhu Chen, Baiwen Chen, Jingyu Li, and others from institutions such as Shanghai Jiao Tong University School of Medicine, Tongji University, and Chongqing Medical University, among others. The work was published on November 13, 2024, in Cellular & Molecular Immunology, with Hua-bing Li as the corresponding author.
Research Flow and Findings
1. Increased Expression of NUDT21 in Inflammatory Diseases
To investigate the role of NUDT21 in inflammatory diseases, the researchers first analyzed mRNA expression profiles in various immune-mediated inflammatory diseases (IMIDs) using the GEO database. The findings revealed that NUDT21 mRNA expression was significantly elevated in inflamed tissues from patients with IBD, psoriasis, RA, and sepsis. Moreover, NUDT21 expression was notably upregulated in bone marrow-derived macrophages (BMDMs) stimulated with IFN-γ/LPS, at both mRNA and protein levels. These results suggest that NUDT21 may play an important role in inflammation progression.
2. Protective Role of NUDT21-Deficient Macrophages Against Colitis
To study the function of NUDT21 in macrophages, the researchers created myeloid-specific NUDT21-deficient mice (NUDT21fl/fl LysMcre, referred to as NUDT21-CKO). They validated the efficient depletion of NUDT21 in BMDMs from CKO mice using RT-qPCR and immunoblot analysis. Subsequently, they induced colitis in mice using a 3% dextran sulfate sodium (DSS) protocol. Compared to wild-type (WT) mice, NUDT21-CKO mice exhibited less body weight loss, reduced colon shortening, and alleviated colon inflammation. These findings suggest that NUDT21 deficiency in macrophages provides protective effects against colitis.
3. NUDT21-Deficient Macrophages Alleviate Hyperinflammatory Responses
To further evaluate the role of NUDT21-deficient macrophages in acute hyperinflammatory conditions, the researchers induced HLH in mice using poly(I:C) and LPS. NUDT21-CKO mice showed significantly improved survival rates, less decline in rectal temperature and body weight, as well as reduced splenic damage scores compared to WT mice. Additionally, cytokine analyses revealed lower mRNA expression of pro-inflammatory cytokines (IL6 and IL1β) in the spleen and liver tissues of CKO mice. Using clodronate liposomes to deplete macrophages, the researchers confirmed that the anti-inflammatory effects were primarily driven by NUDT21-deficient macrophages.
4. Decreased Proinflammatory Traits in NUDT21-Deficient Macrophages
In HLH models, NUDT21-deficient macrophages exhibited increased cell viability with reduced levels of pro-inflammatory cytokines such as IL6 and TNFα, as well as lower macrophage death. Similarly, in DSS colitis models, intestinal macrophages from NUDT21-CKO mice displayed reduced inflammatory cytokine production. These results confirm that NUDT21 deficiency reduces proinflammatory traits in macrophages.
5. Enhanced Autophagic Activity in NUDT21-Deficient Macrophages
To investigate the molecular basis of the anti-inflammatory effects, the researchers performed high-throughput RNA-seq on BMDMs from WT and NUDT21-CKO mice. They identified 2,707 differentially expressed genes, with gene enrichment analyses revealing significant effects on autophagy-related pathways. Transmission electron microscopy (TEM) and immunostaining demonstrated increased autophagic vacuoles and enhanced autophagic activity in NUDT21-deficient macrophages.
Further analysis showed that NUDT21-deficient CKO macrophages exhibited increased expression and mRNA stability of essential autophagy-related genes such as MAP1LC3B and ULK2.
6. NUDT21 Modulates the mRNA Stability of Autophagy Genes
NUDT21 binds specifically to UGUA motifs to regulate distal PAS usage, and its absence results in shortened 3’UTRs. Using the DAPARS algorithm, the researchers observed widespread 3’UTR shortening in NUDT21-CKO macrophages. This resulted in increased mRNA stability of important autophagy genes, enhancing autophagic activity and reducing NF-κB signaling. Consequently, inflammatory cytokine production was decreased.
Conclusion and Significance
This study revealed that NUDT21 regulates autophagic activity in macrophages during inflammation by modulating the 3’UTR length of key autophagy genes. NUDT21-deficient macrophages demonstrated enhanced autophagy and reduced inflammatory cytokine secretion, leading to protective effects in colitis and HLH models. This finding highlights the complex interplay between RNA metabolism and autophagy regulation, providing new insights into inflammation.
Study Highlights
- NUDT21 is significantly upregulated in various inflammatory diseases, emphasizing its role in inflammation progression.
- NUDT21-deficient macrophages exhibit enhanced autophagic activity and lower pro-inflammatory cytokine secretion, providing protection against inflammatory conditions.
- NUDT21 modulates 3’UTR length of mRNAs, affecting mRNA stability of autophagy-related genes such as MAP1LC3B and ULK2.
Research Value
By elucidating the role of NUDT21 in macrophage-mediated inflammation, this study identifies potential therapeutic targets for inflammatory diseases. Targeting NUDT21 or associated 3’UTRs in autophagy genes could enable precise interventions to modulate inflammation without impairing overall macrophage functionality, paving the way for advanced anti-inflammatory therapies with clinical potential.