Viral miRNA Antagomirs Rescue Antigen Presentation Pathway in Ocular HSV-1 Infection

Herpes simplex virus type 1 (HSV-1) establishes lifelong latency by deploying multiple mechanisms to evade host immunity. However, the role of virus-encoded microRNAs (v-miRs) in modulating antiviral immune responses remains incompletely understood. We previously showed that topical inhibition of the HSV-1 v-miRs miR-H6-3p and miR-H27 in infected animals reduces viral titers and restores immune homeostasis. Here, using a murine model of corneal HSV-1 infection, we examined whether in vivo inhibition of these v-miRs restores expression of genes involved in antigen processing and presentation. Targeted transcript profiling revealed that inhibition of miR-H6-3p altered 25 genes (19 upregulated, 6 downregulated), whereas inhibition of miR-H27 affected 20 genes (10 upregulated, 10 downregulated) relative to scramble controls. To determine the functional relevance of these findings in human antigen-presenting cells, we next examined v-miR activity in primary human macrophages. Overexpression of miR-H6-3p or miR-H27 mimics significantly impaired antigen processing of BODIPY-conjugated ovalbumin and reduced CD4 + T-cell proliferation in macrophage-T cell coculture assays, consistent with defective antigen presentation. In silico analysis identified predicted v-miR binding sites in the 3 ′ UTRs of ERAP1, TAP2, and CD2. Dual-luciferase assays confirmed direct targeting by miR-H6-3p and miR-H27, accompanied by reduced transcript abundance of these genes in v-miR-transfected cells. Collectively, these findings reveal a previously unrecognized mechanism of HSV-1 immune evasion in which viral microRNAs suppress antigen-processing and presentation pathways. Targeting v-miRs may therefore represent a therapeutic strategy to restore antigen presentation and antiviral immunity in the cornea. • Inhibition of HSV-1 encoded microRNAs (miR-H6-3p and miR-H27) upregulates antigen processing and presentation genes. • miR-H6-3p and miR-H27 overexpression attenuate antigen processing. • Antigen processing genes ERAP1, TAP2, CD2 are targeted by miR-H6-3p and miR-H27 • Luciferase assays confirm direct HSV-1 encoded microRNAs binding to ERAP1/TAP2/CD2 3'UTRs. • Inhibition of viral microRNA can be a promising therapeutic strategy for immune modulation during viral infection.