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HuR promotes miRNA-mediated upregulation of NFI-A protein expression in MDSCs during murine sepsis
Isatou Bah1, Tuqa Alkhateeb1, Ajinkya Kumbhare1
1Department of Internal Medicine, East Tennessee State University College of Medicine, Johnson City, TN, 37614, USA.
Abstract:
Myeloid-derived suppressor cells (MDSCs) contribute to high mortality rates during sepsis, but how sepsis induces MDSCs is unclear. Previously we reported that microRNA (miR)-21 and miR-181b reprogram MDSCs in septic mice by increasing levels of DNA binding transcription factor, nuclear factor 1 (NFI-A). Here, we provide evidence that miR-21 and miR-181b stabilize NFI-A mRNA and increase NFI-A protein levels by recruiting RNA-binding proteins HuR and Ago1 to its 3' untranslated region (3'UTR). We also find that the NFI-A GU-rich element (GRE)-binding protein CUGBP1 counters miR-21 and miR-181b dependent NFI-A mRNA stabilization and decreases protein production by replacing 3'UTR bound Ago1 with Ago2. We confirmed the miR-21 and miR-181b dependent reprogramming pathway in MDSCs transfected with a luciferase reporter construct containing an NFI-A 3'UTR fragment with point mutations in the miRNA binding sites. These results suggest that targeting NFI-A in MDSCs during sepsis may enhance resistance to uncontrolled infection.
Insights
Sepsis increases myeloid-derived suppressor cells (MDSCs) by stabilizing nuclear factor 1 (NFI-A) mRNA via microRNAs (miRs). Targeting NFI-A in MDSCs may improve sepsis outcomes.
Area of Science:
- Immunology
- Molecular Biology
- Genetics
Background:
- Myeloid-derived suppressor cells (MDSCs) are key drivers of sepsis-induced mortality.
- The precise mechanisms by which sepsis induces MDSCs remain incompletely understood.
- Previous work identified microRNA (miR)-21 and miR-181b as critical in reprogramming MDSCs during sepsis by upregulating nuclear factor 1 (NFI-A).
Purpose of the Study:
- To elucidate the molecular mechanisms underlying miR-21 and miR-181b-mediated NFI-A regulation in MDSCs during sepsis.
- To investigate the roles of RNA-binding proteins HuR, Ago1, Ago2, and CUGBP1 in this pathway.
- To validate the functional significance of the identified miRNA-NFI-A regulatory axis.
Main Methods:
- Luciferase reporter assays utilizing constructs with wild-type and mutated NFI-A 3' untranslated regions (3'UTRs).
- Analysis of RNA-binding protein recruitment (HuR, Ago1, Ago2) to the NFI-A 3'UTR.
- Investigation of CUGBP1's role in modulating miRNA effects on NFI-A mRNA and protein.
- Experimental validation in MDSCs.
Main Results:
- miR-21 and miR-181b enhance NFI-A protein levels by recruiting HuR and Ago1 to the NFI-A 3'UTR, stabilizing its mRNA.
- CUGBP1 antagonizes this process by displacing Ago1 with Ago2 at the NFI-A 3'UTR, thereby reducing NFI-A protein production.
- Mutational analysis of miRNA binding sites within the NFI-A 3'UTR confirmed the regulatory pathway.
Conclusions:
- Sepsis-induced MDSC reprogramming involves a complex interplay of specific miRNAs and RNA-binding proteins regulating NFI-A.
- The findings reveal a novel mechanism of post-transcriptional gene regulation critical for MDSC function in sepsis.
- Targeting NFI-A within MDSCs presents a potential therapeutic strategy to mitigate sepsis severity and improve patient outcomes.
