Retroviral integration is a crucial step in the viral life cycle, yet its impact on chromatin stability remains unclear. We hypothesize that prototype foamy virus (PFV) integration leads to nucleosome disassembly. To test this, we use dual-beam optical tweezers to perform real-time, single-molecule measurements of PFV integration into nucleosomal substrates. Both constant-force and force-ramp assays examine how PFV alters nucleosome mechanics. Destabilization is monitored through increased DNA extension under constant force and changes in characteristic outer turn unwrapping forces. We compare integration outcomes with different cofactors (Mg 2+ vs. Ca 2+ ) to assess how cofactor choice influences the integration process. Complementary atomic force microscopy and fluorescence-force microscopy provide additional insight. Preliminary force-extension studies on plasmid DNA reveal distinct cofactor-dependent integration mechanisms—Mg 2+ promotes strand cutting events, while Ca 2+ produces altered hysteresis patterns. These contrasting mechanical signatures suggest that the integration cofactor affects the hypothesized disassembly. Future directions include force ramp and constant force assays of PFV integration to nucleosomal DNA in three different groups—no cofactor, Mg 2+ , and Ca 2+ . Differences in the stretching dynamics would be used to infer the structural influences of PFV on nucleosomal DNA in the presence of each cofactor. PFV’s well-characterized integration mechanism makes it an ideal model system for studying virus-chromatin interactions, with implications for understanding pathogenic retroviruses like HIV. Its non-pathogenicity gives it high potential as a vector in gene therapy, as well.
Sara A.A. Mohamed (Sun,) studied this question.