Regulation and function of the HPV16 CircE7 RNA
9 affiliations
- 1Department of Dermatology, University of Texas Southwestern Medical Center (UTSW), Dallas, Texas, USA
- 2Division of Hematology-Oncology, Department of Internal Medicine, Chang Gung Memorial Hospital Linkou, Linkou, Taoyuan, Taiwan
- 3Medical School, UTSW, Dallas, Texas, USA
- 4Department of Dermatology, University of Colorado Anschutz Medical Campus (CU AMC), School of Medicine, Aurora, Colorado, USA
- 5Medical School, Louisiana State University Health Sciences Center, New Orleans, Louisiana, USA
- 6Oral and Craniofacial Molecular Biology, Virginia Commonwealth University, Richmond, Virginia, USA
- 7Department of Pathology, UTSW, Dallas, Texas, USA
- 8Department of Otolaryngology, UTSW, Dallas, Texas, USA
- 9University of Colorado Cancer Center, Aurora, Colorado, USA
Abstract
High-risk human papillomaviruses (HPV), including HPV16, produce circular RNA that encompasses the E7 oncogene (circE7). CircE7 can be detected in HPV16-positive cells and tumors, is preferentially localized to the cytoplasm, is N6-methyladenosine (m6A)-modified, and can be translated into the E7 oncoprotein. Here, we explored the regulation and function of circE7. Mutation of m6A motifs flanking the backsplice junction revealed a single m6A motif to be essential for circE7 formation. Mutation of this m6A motif promoted linear splicing of the E6*I splice site (226^409), suggesting that linear and circular E7 splicing are inversely regulated. Additionally, mutation of an IRES-like motif in circE7 significantly decreased E7 protein expression, without having significant effects on circE7 RNA levels. Knockdown of YTHDC1, but not other m6A-binding proteins, decreased both circE7 RNA and protein expression. BaseScope ISH was used to confirm the expression of circE7 in head and neck squamous cell carcinoma cell lines and tumors. Using both qRT-PCR and BaseScope ISH, we found that serum and amino acid starvation significantly increased circE7. Finally, we generated an HPV16 genome with mutations in the circE7 m6A motif (Mut2). Stable transduction of primary keratinocytes with Mut2 confirmed the loss of circE7 and increased expression of E6*I. The Mut2 HPV16 genome exhibited significantly decreased viral replication but an increased ability to transform primary keratinocytes. Our studies reveal that the precise regulation of circE7 and E6*I by m6A is critical for the ability of HPV16 to infect and transform keratinocytes.
IMPORTANCE: High-risk human papillomaviruses (HPVs), such as HPV16, must carefully control how much E6 and E7 proteins they make. This study shows that HPV16 toggles a single chemical tag on the viral RNA (an m⁶A mark) to control the production of early region RNAs, including a circular RNA called circE7. The same site coordinately regulates splicing of the E6*I isoform. CircE7 uses m⁶A-binding proteins to control its production and a specific sequence to promote its translation. It is present in HPV-positive cancers and can respond to nutrient starvation. Regulation of circE7 through this m6A site also impacted viral replication and transformation capacity, indicating that this regulatory mechanism is critical for HPV biology.
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BibTeX
@article{lee2026regulation,
title = {{Regulation and function of the HPV16 CircE7 RNA}},
author = {Eunice E. Lee and Yihui Huang and Eleanor Dowell and Kaitlyn Walsh and Calvin Hosler and Austin Witt and Odera Ifeacho and Doreen Palsgrove and Andrew Day and Claire James and Richard C. Wang},
journal = {Journal of Virology},
year = {2026},
month = sep,
doi = {10.1128/jvi.01284-26},
url = {https://doi.org/10.1128/jvi.01284-26},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Regulation and function of the HPV16 CircE7 RNA
AU - Eunice E. Lee
AU - Yihui Huang
AU - Eleanor Dowell
AU - Kaitlyn Walsh
AU - Calvin Hosler
AU - Austin Witt
AU - Odera Ifeacho
AU - Doreen Palsgrove
AU - Andrew Day
AU - Claire James
AU - Richard C. Wang
PY - 2026
DA - 2026/09/17
JO - Journal of Virology
DO - 10.1128/jvi.01284-26
UR - https://doi.org/10.1128/jvi.01284-26
AB - High-risk human papillomaviruses (HPV), including HPV16, produce circular RNA that encompasses the E7 oncogene (circE7). CircE7 can be detected in HPV16-positive cells and tumors, is preferentially localized to the cytoplasm, is N6-methyladenosine (m6A)-modified, and can be translated into the E7 oncoprotein. Here, we explored the regulation and function of circE7. Mutation of m6A motifs flanking the backsplice junction revealed a single m6A motif to be essential for circE7 formation. Mutation of this m6A motif promoted linear splicing of the E6*I splice site (226^409), suggesting that linear and circular E7 splicing are inversely regulated. Additionally, mutation of an IRES-like motif in circE7 significantly decreased E7 protein expression, without having significant effects on circE7 RNA levels. Knockdown of YTHDC1, but not other m6A-binding proteins, decreased both circE7 RNA and protein expression. BaseScope ISH was used to confirm the expression of circE7 in head and neck squamous cell carcinoma cell lines and tumors. Using both qRT-PCR and BaseScope ISH, we found that serum and amino acid starvation significantly increased circE7. Finally, we generated an HPV16 genome with mutations in the circE7 m6A motif (Mut2). Stable transduction of primary keratinocytes with Mut2 confirmed the loss of circE7 and increased expression of E6*I. The Mut2 HPV16 genome exhibited significantly decreased viral replication but an increased ability to transform primary keratinocytes. Our studies reveal that the precise regulation of circE7 and E6*I by m6A is critical for the ability of HPV16 to infect and transform keratinocytes. IMPORTANCE: High-risk human papillomaviruses (HPVs), such as HPV16, must carefully control how much E6 and E7 proteins they make. This study shows that HPV16 toggles a single chemical tag on the viral RNA (an m⁶A mark) to control the production of early region RNAs, including a circular RNA called circE7. The same site coordinately regulates splicing of the E6*I isoform. CircE7 uses m⁶A-binding proteins to control its production and a specific sequence to promote its translation. It is present in HPV-positive cancers and can respond to nutrient starvation. Regulation of circE7 through this m6A site also impacted viral replication and transformation capacity, indicating that this regulatory mechanism is critical for HPV biology.
SN - 0022-538X
SN - 1098-5514
ER -