Single-Cell Genomic Profiling Identifies an H3F3B–SEPTIN6 Axis Associated with Malignant Epithelial Phenotypes in Lung Adenocarcinoma

Abstract Background Lung adenocarcinoma is an important cause of cancer morbidity and mortality among women, yet sex-specific molecular features of malignant epithelial populations remain incompletely understood. This study investigated the expression and functional significance of H3F3B and its association with SEPTIN6 in LUAD. Methods Single-cell RNA sequencing data from three publicly available lung datasets were integrated to characterize epithelial cell populations and identify genes associated with malignant phenotypes. Differential expression, pathway enrichment, protein–protein interaction, and gene co-expression analyses were performed to characterize H3F3B-associated transcriptional programs. H3F3B expression was further evaluated in three paired LUAD and adjacent tissue specimens by reverse transcription quantitative polymerase chain reaction, Western blotting, and immunohistochemistry. Functional effects of H3F3B were investigated in A549 cells using overexpression and siRNA-mediated knockdown, followed by proliferation, migration, and invasion assays. SEPTIN6 expression was assessed following H3F3B perturbation. Results Integration of 665,761 epithelial cells from 878 samples identified distinct epithelial populations, including a prominent Secretory Club/AT2-like malignant population. H3F3B was significantly upregulated in malignant epithelial cells and showed increased expression across LUAD stages. H3F3B expression was positively correlated with SEPTIN6 expression in the integrated dataset. In paired patient tissues, H3F3B expression was higher in LUAD tissues than in adjacent tissues at both the mRNA and protein levels. H3F3B overexpression increased A549 cell proliferation, migration, and invasion, whereas H3F3B knockdown produced the opposite effects. SEPTIN6 expression increased following H3F3B overexpression and decreased following H3F3B knockdown. Conclusion H3F3B is associated with malignant epithelial states in LUAD and promotes proliferative, migratory, and invasive phenotypes in A549 cells. Its positive association with SEPTIN6 and the concordant regulation of SEPTIN6 following H3F3B perturbation suggest that the H3F3B–SEPTIN6 pathway may contribute to LUAD progression. Further studies are required to define the molecular mechanism underlying this regulatory relationship and to establish its clinical relevance.