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 downregulated genes ERAP1, TAP2, and CD2. Dual-luciferase assays confirmed direct targeting of key antigen processing and presentation genes by miR-H6-3p and miR-H27, accompanied by reduced transcript abundance 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.