HAdV55 reprograms host 3D genome architecture and mitochondrial metabolism to drive pathogenesis.

Human adenovirus type 55 (HAdV55), an emerging recombinant pathogen linked to severe respiratory outbreaks, exhibits heightened virulence and replication efficiency, yet its mechanisms of host interaction remain poorly resolved. Here, we integrate transcriptomic and 3D genomic analyses to define the spatiotemporal interplay between HAdV55 and A549 cells. Temporal profiling revealed triphasic host transcriptional reprogramming, with distinct clusters of genes governing cell survival, apoptosis, and innate immunity dynamically modulated during infection. Crucially, HAdV55 reshapes the 3D chromatin architecture, inducing compartmental shifts on chromosome 15 (A-to-B transitions at 72-96 h) and selectively localizing its genome to transcriptionally active chromatin regions. Strikingly, the viral genome exhibits progressive interactions with mitochondrial energy metabolism genes (ATP6, ND1, CYB), suggesting mitochondrial hijacking to fuel replication. Functional validation via gene silencing identified ABL2, FANCI, STK4, TRIM24, TRIM2 and MAPK10 as critical host regulators restricting viral proliferation, with knockdowns amplifying viral loads over to 15-fold. This study also establishes, for the first time, a spatial link between adenoviral infection and mitochondrial genome dynamics, offering a paradigm for investigating virus-host interactions in three-dimensional genomic contexts.