Human papillomaviruses (HPVs) are DNA viruses with epithelial tropism. High-risk types of HPV are the causative agents of the majority of cervical cancers and are responsible for a number of other anogenital as well as oropharyngeal cancers. The life cycle of HPV is closely linked to the differentiation state of its host cell and is dependent on the activation of specific pathways of the DNA damage response. Several proteins from the ataxia telangiectasia mutated and the ataxia telangiectasia mutated and Rad3-related DNA repair pathways, which are essential for maintaining genomic stability in cells, are upregulated in HPV-positive cells and are required for viral replication. Our studies examine the expression of 5 such DNA repair factors-pCHK2, pCHK1, FANCD2, BRCA1, and H2AX-in cervical specimens from patients diagnosed with low-grade, intermediate-grade, or high-grade lesions. The percentage of cells expressing pCHK2, pCHK1, FANCD2, and BRCA1 is significantly higher in high-grade squamous intraepithelial lesions compared with that of either low-grade squamous intraepithelial lesions or normal tissue, particularly in differentiated cell layers. In addition, the distribution of this staining throughout the epithelium is altered with increasing lesion grade. This study characterizes the expression of pCHK2, pCHK1, FANCD2, H2AX and BRCA1 during cervical cancer progression and provides additional insight into the role of these DNA damage response proteins in viral transformation.