Virus–Host Protein–Protein Interactions between Human Papillomavirus 16 E6 A1 and D2/D3 Sub-Lineages: Variances and Similarities

Abstract

High-risk strains of human papillomavirus are causative agents for cervical and other mucosal cancers, with type 16 being the most frequent. Compared to the European Prototype (EP; A1), the Asian-American (AA; D2/D3) sub-lineage seems to have increased abilities to promote carcinogenesis. Here, we studied protein–protein interactions (PPIs) between host proteins and sub-lineages of the key transforming E6 protein. We transduced human keratinocyte with EP or AA E6 genes and co-immunoprecipitated E6 proteins along with interacting cellular proteins to detect virus–host binding partners. AAE6 and EPE6 may have unique PPIs with host cellular proteins, conferring gain or loss of function and resulting in varied abilities to promote carcinogenesis. Using liquid chromatography-mass spectrometry and stringent interactor selection criteria based on the number of peptides, we identified 25 candidates: 6 unique to AAE6 and EPE6, along with 13 E6 targets common to both. A novel approach based on pathway selection discovered 171 target proteins: 90 unique AAE6 and 61 unique EPE6 along with 20 common E6 targets. Interpretations were made using databases, such as UniProt, BioGRID, and Reactome. Detected E6 targets were differentially implicated in important hallmarks of cancer: deregulating Notch signaling, energetics and hypoxia, DNA replication and repair, and immune response.

Targeting the Human Papillomavirus 16 E6 Oncoprotein with Antibodies

“…This could indicate that these candidates have limited binding affinity to the endogenous E6 protein. Based on our results with C26, such sdAbs likely bind the folded E6 protein but E6 is also known to interact with 50 or more host proteins (summarized in Dayer et al 2020 with references therein) using residues distributed throughout the protein. This means that any molecule targeting E6 must compete with host proteins.…”

Metabolic Reprogramming in Cancer: Role of HPV 16 Variants

“…Furthermore, the Wnt/β-catenin pathway has been shown to increase the Warburg effect by suppressing mitochondrial respiration by reducing cytochrome oxidase transcription, an essential enzyme for OXPHOS [154]. On the other hand, AA sublineage variants increase mTOR activation, which, like the Wnt/β-catenin pathway, leads to energy metabolism activation through HIF-1α [13]. The E6 variant E-A126 with the R8Q mutation promotes the glycolytic pathway by activating the Wnt/β-catenin pathway.…”

“…One of the ways that could also be participating in the positive regulation of glycolysis through the AA sublineage variant is by binding to the gamma-2 subunit of the protein kinase activated by 5'-AMP On the other hand, the Asian variant of E6, containing the amino acid change D25E in the E6 oncoprotein, negatively regulates the expression of the AIFM2 protein [16], known as apoptosis-inducing factor mitochondria associated 2, which contains a domain oxidoreductase NADH [166]. It has also been observed that the variant of the AA sublineage binds to cytochrome C [13] and could be interfering at levels such as the Asian variant D25E [13,16,167], which also increases the expression levels of ENO1 (Figure 4) [16]. The low levels of AIFM2 and cytochrome C, as well as the increase in the expression of ENO1 induced by the variant AA and As, may affect the reduction of oxidative phosphorylation, favoring the production of energy through aerobic glycolysis, one of the main characteristics of metabolic reprogramming in cancer.…”

“…This suggests that the expression of CAIX is directly related to the oncogenic potential of the AA sublineage variants of the E6 oncoprotein, where there is a more significant increase compared to the E-prototype [ 163 ]. One of the ways that could also be participating in the positive regulation of glycolysis through the AA sublineage variant is by binding to the gamma-2 subunit of the protein kinase activated by 5’-AMP (PRKAG2) [ 13 ], a regulator key to vigorous metabolism that activates energy production pathways and inhibits energy-consuming processes [ 13 ].…”

“…Currently, several reports show that human papillomavirus (HPV) 16 variants alter the expression of various genes related to apoptosis, adhesion, metastasis, angiogenesis, and metabolism [ 12 ]. These variants interact with multiple cellular proteins such as p53, retinoblastoma protein (pRb), p300/CBP, and pyruvate kinase 2 (PKM2), promoting gain or loss of their functions [ 13 ]. These have also been associated with a more significant oncogenic potential of these variants and the severity of precancerous lesions [ 14 , 15 ].…”