Virus–Host Interaction: Investigating Novel Transcription Factors Involved in Coupling Human Papillomavirus Life Cycle and Epithelial Differentiation

High-risk human papillomavirus (HPV) is the etiological agent of nearly all cervical cancers, with HPV-18 being the second most prevalent type. The HPV life cycle is closely associated with epithelial differentiation, a process governed by cellular transcription factors (TFs). During progression toward malignancy, HPV disrupts differentiation and promotes uncontrolled cell proliferation. We aimed to identify differentiation-modulated TFs linking viral transcription to differentiation, providing insights into mechanisms driving viral life cycle and pathogenesis. DNA-binding activity of 345 TFs was compared between undifferentiated and differentiated keratinocytes to identify differentiation-associated TFs. In silico analyses identified putative binding sites for these TFs within the HPV-18 long control region (LCR). Chromatin immunoprecipitation confirmed direct binding of PAX6, HMGB1, and NFE2 to the LCR, but not FOXI1. Immunohistochemistry in keratinocyte raft cultures demonstrated differentiation-dependent expression patterns for all four TFs. Functional assays revealed that each TF is capable of modulate HPV-18 early promoter activity, with effects varying by differentiation status. Notably, expression of these TFs was disrupted by the viral oncoproteins E6 and E7, underscoring a mechanism by which HPV alters host differentiation. These findings identify novel differentiation-linked regulators of HPV-18 transcription and highlight host targets exploited by the virus in its life cycle and pathogenesis.