Characterization of Stress Responsive MicroRNAs and Their Roles in T Cell Development

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2014-01-06

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Belkaya, Serkan

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Abstract

Physiological stress evokes rapid changes in both the innate and adaptive immune responses. Immature αβ T cells developing in the thymus are particularly sensitive to stress, with infections and/or exposure to lipopolysaccharide or glucocorticoids eliciting a rapid apoptotic program. MicroRNAs (miRs) are short, non-coding RNAs that play critical roles in the immune system by targeting diverse mRNAs. We hypothesized that a subset of thymically encoded miRs would be stress responsive and modulate thymopoiesis. Thymic miR profiling revealed 18 distinct miRs that are dysregulated more than 1.5-fold in response to lipopolysaccharide or the synthetic glucocorticoid dexamethasone. These stress-responsive miRs are dynamically regulated in distinct thymocyte subsets. We utilized both transgenic and gene-targeting approaches to study the impact of these miRs on thymopoiesis under normal and stress conditions. MiR-181d is the most down-regulated thymic miR in response to stress. The over-expression of miR-181d in developing thymocytes reduced the number of immature CD4+CD8+ thymocytes. Lipopolysaccharide or dexamethasone injections caused a 4-fold greater loss of these cells than in the wild type controls. The targeted elimination of miR-181d resulted in a thymus stress-responsiveness similar to wild-type mice, suggesting a functional redundancy between miR-181 family members. Gene expression comparisons further indicated that miR-181d affects a number of stress, metabolic, and signaling pathways. These findings demonstrate that selected miRs enhance stress-mediated thymic involution in vivo. MiR-185, another stress-responsive miR in murine thymus, is haploinsufficient in almost all individuals with 22q11.2 deletion/DiGeorge syndrome that can present with immune, cardiac, parathyroid, and psychological problems. The molecular targets of miR-185 in thymocytes are unknown. Transgenic expression of miR-185 attenuated thymopoiesis at the TCRβ-selection checkpoint and during positive selection. This caused a peripheral T cell lymphopenia. Mzb1, NFATc3, and Camk4 were identified as novel miR-185 targets. Elevations in miR-185 enhanced TCR-dependent intracellular calcium levels, while a knockdown of miR-185 diminished these calcium responses. These effects concur with reductions in Mzb1, an endoplasmic reticulum calcium regulator. Consistent with the haploinsufficiency of miR-185, Mzb1 levels were elevated in thymocyte extracts from several 22q11.2 deletion/DiGeorge syndrome patients. These findings indicate that miR-185 regulates T cell development through its targeting of several mRNAs including Mzb1.

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