人乳头瘤病毒（HPVs）与约5%-10%的人类癌症相关，尤其是几乎99%的宫颈癌。HPV在病毒生命周期中与分层上皮（分化层）交互的机制，以及致癌机制仍不清楚。本研究采用单细胞转录组分析研究了HPV阳性宫颈癌组织的病毒基因和宿主细胞分化相关的异质性。我们检查了HPV16的基因E1、E6和E7，并发现它们在九个上皮细胞簇中表达不同。我们发现三个上皮细胞簇中HPV阳性细胞的比例最高（分别为33.6%、37.5%和32.4%），而另外两个上皮细胞簇中的比例最低（分别为7.21%和5.63%）。值得注意的是，含有最多HPV阳性细胞的簇与正常上皮层标记显著偏离，表现出功能异质性和上皮结构改变，表明癌组织中存在显著的分子异质性，这些细胞与正常上皮细胞相比具有独特/不同的基因特征。与HPV阴性细胞相比，这些HPV阳性细胞在细胞外基质、细胞粘附、增殖和凋亡等方面的基因表达不同。此外，病毒致癌基因E6和E7似乎通过不同的途径改变上皮功能，从而促进宫颈癌的发展。我们从分层异质性的新视角研究了HPV和宿主基因的转录组。我们的结果表明，在上皮细胞簇中HPV的表达存在差异，并且与病毒致癌基因相关的上皮异质性为这一关键研究领域提供了生物学洞见。© 2023 The Authors. Journal of Medical Virology published by Wiley Periodicals LLC.

Human Papillomaviruses (HPVs) are associated with around 5%-10% of human cancer, notably nearly 99% of cervical cancer. The mechanisms HPV interacts with stratified epithelium (differentiated layers) during the viral life cycle, and oncogenesis remain unclear. In this study, we used single-cell transcriptome analysis to study viral gene and host cell differentiation-associated heterogeneity of HPV-positive cervical cancer tissue. We examined the HPV16 genes-E1, E6, and E7, and found they expressed differently across nine epithelial clusters. We found that three epithelial clusters had the highest proportion of HPV-positive cells (33.6%, 37.5%, and 32.4%, respectively), while two exhibited the lowest proportions (7.21% and 5.63%, respectively). Notably, the cluster with the most HPV-positive cells deviated significantly from normal epithelial layer markers, exhibiting functional heterogeneity and altered epithelial structuring, indicating that significant molecular heterogeneity existed in cancer tissues and that these cells exhibited unique/different gene signatures compared with normal epithelial cells. These HPV-positive cells, compared to HPV-negative, showed different gene expressions related to the extracellular matrix, cell adhesion, proliferation, and apoptosis. Further, the viral oncogenes E6 and E7 appeared to modify epithelial function via distinct pathways, thus contributing to cervical cancer progression. We investigated the HPV and host transcripts from a novel viewpoint focusing on layer heterogeneity. Our results indicated varied HPV expression across epithelial clusters and epithelial heterogeneity associated with viral oncogenes, contributing biological insights to this critical field of study.© 2023 The Authors. Journal of Medical Virology published by Wiley Periodicals LLC.