MicroRNA-451 Regulates T Helper Cell Responses

Thesis (Ph.D.)--University of Rochester. School of Medicine & Dentistry. Dept. of Translational Biomedical Sciences, 2015.Background: Malaria parasite evasion of host defense mechanisms leads to uncontrolled parasite growth, anemia, morbidity, and often mortality. A number of studies have shown that miR-451 may be a key regulator of immune function. Down regulation of miR-451 promotes the release of proinflammatory cytokines from dendritic cells, and NOTCH1 mediated repression of miR-451 promotes Tcell proliferation in a mouse leukemia model. The goal of this study is to define the underlying genetic and molecular mechanisms that lead to increased host immune responses in miR-451-/- mice during malaria infection. Results: Studies by our lab using the uncomplicated malaria mouse model of Plasmodium yoelii (P. yoelii) infection demonstrated that mice lacking miR-451 (miR-451-/-) have greatly accelerated parasite clearance compared to wild type (WT) controls. The accelerated parasite clearance is dependent on increased CD4+ T-cell proliferation responses to infection. Using RNAseq analysis we found 5028 genes were differentially expressed between WT and miR-451-/- CD4+ T cells post infection (55 genes with a log2-fold change > 2) while only 88 genes differed pre-infection. Western blot and quantitative real time PCR analysis identified increased Myc expression in miR-451-/- CD4+ T cells post infection. Myc has a key role in promoting CD4+ T-cell proliferation and our studies demonstrated that miR-451 has a negative effect CD4+ T-cell proliferation that is at least in part Myc dependent. MiR-451-/- CD4+ T cells have increased cell proliferation when stimulated in vitro compared to WT CD4+ T cells. In addition, antagomir inhibition of miR-451 in WT T cells causes an increase in WT CD4+ Tcell proliferation post stimulation in vitro. We have also shown that pharmacologic inhibition of c-myc blunts miR-451-/- CD4+ T-cell proliferation in vitro. Taken together these data indicate that miR-451 is a major regulator of CD4+ T-cell proliferation, in part via regulation of Myc expression post T-cell activation. Discussion: Our study is the first to demonstrate that the loss of a miRNA generates a protective host immune response against malaria infection. Mice lacking miR-451 have an enhanced CD4+ T-cell proliferative response post malaria infection. Decreased expression of miR-451 promotes T helper proliferation in part as a result of increased Myc expression. Among the 5028 differentially expressed genes, we have identified candidate that may be involved in cell proliferation and cytokine pathways (ie: NFKB pathway genes: Psmb8 [1.55 log2 fold change] and Ikbkb [0.72 log2 fold change]). We plan to confirm these pathways using bioinformatics analysis. This study has many clinical implications, including treating patients with miR-451 antagomirs or an adoptive cell therapy strategy in which miR-451 expression in antigen specific CD4+ T cells is blunted, and modified cells are reintroduced into the patient to enhance the CD4+ T-cell responses to malaria infection. It is also relevant to many other diseases in which T-cell proliferation is part of the pathogenesis. For example, we found in our heart transplant mouse model that miR-451-/- heart recipients had accelerated CD4+ T cell mediated transplant rejection compared to WT recipients. Altering miR-451 expression may be therapeutically beneficial in either limiting or increasing T-cell responses