Early evidence for an alternative mode of DNA replication

Evidence for a rolling circle mode of DNA replication for bovine papillomavirus type 1 (BPV-1)

(electron micrograph image from Dasgupta et. al. "Rolling-circle replication of a high-copy BPV-1 plasmid" (Journal of Molecular Biology 228(1), 1992, pp.1-6) reproduced with the permission of the publisher).

Project Background & Introduction

We were stimulated to resurrect our study of papillomavirus rolling circle replication (RCR) after an interval of over 25 years as a subject too long neglected both by ourselves and possibly by the wider papillomavirus research community which had concluded in the intervening years that, at least for human papillomaviruses (HPVs), recombination-dependent replication (RDR) was the primary mechanism of viral genome amplification in the vegetative phase of the papillomavirus replication cycle. Knowledge of the mechanism of RCR in other virus and phage systems had also come a long way since the seminal breakthrough by Ilyina and Koonin, published in 1992, which identified specific evolutionarily conserved amino acid sequence motifs common to the replication initiation factors encoded by a wide range of extrachromosomal circular DNA molecules. Furthermore, due in large part to the development of rapid DNA sequencing technology in the modern era, a vast number of new human and animal papillomavirus types had been sequenced and were thus available for comparative analysis.

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Predicted Latent Endonuclease Function of Papillomavirus E1 Genes: Duplication of an Endonuclease Domain in the Evolution of the Papillomavirus E1 Gene is Suggested by Divergent Localizations of RCR Initiator Motifs in the BPV-1 and HPV-16 E1 Proteins

Contributors

Stan Burnett

Santanu Dasgupta

Goal

It has long been believed that the papillomavirus E1 gene possesses just a single enzymatic activity conferred by its C-terminal helicase domain. However, we have recently identified amino acid sequence motifs characteristic of the HUH endonuclease superfamily of rolling circle replication (RCR) initiators within the E1 proteins of BPV-1 and closely related deltapapillomaviruses (BPV-2, BPV-13, and BgPV-1). Putative Motif II (HLH/HLQ) and Motif III (YDHKY) sequence elements were identified precisely bordering their E1 DNA binding domains (DBDs). A potential Motif I sequence (VLTPLQ) was also identified in BPV-1 just proximal to the Motif II sequence. RCR initiator motifs were also identified in the HPV-16 E1 gene, but unlike those of BPV-1, which lie outside of the minimal DBD, the HPV-16 motifs II and III are situated within the E1 DBD in close proximity to one another within the folded protein, resembling the situation in all known HUH endonucleases. Our working hypothesis is that there was a duplication of an endonuclease domain and subsequent divergence in the mechanism of regulation of an endonuclease function between different papillomaviruses, as exemplified by BPV-1 and HPV-16. FROM 1st April 2023 FOLLOW PROGRESS OF THIS PROJECT AT www.StanleyBurnett.com

Hypothesis

Bovine papillomavirus type 1 (BPV-1) and human papillomavirus type 16 (HPV-16) E1 genes encode rolling-circle replication initiator proteins.

Experiment findings

Experiment findings that have been added to share the progress of the project.

List of experiment findings from the project

Final papers

Preprints and articles that have been added as the result of this project.

List of final papers from the project

References

Research that was mentioned or added to this project.

List of research referenced in this project

Project log

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Project log

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February 2023

Stan Burnett has added an update

Feb 15,2023

Identification of RCR motifs in the HPV-16 E1 protein

Rolling circle replication (RCR) Motif II and Motif III sequence elements HIQ and YWYK were identified in the HPV-16 E1 coding sequence. These elements were situated in the E1 DNA binding domain (DBD) within localised segments of beta-sheet (HIQ) and alpha-helical (YWYK) structure corresponding to the beta-2 and alpha-5 folds (which are conserved structures among the E1 DBDs of papillomaviruses) and are predicted to lie in close proximity within the 3-dimensional folded structure of the E1 protein. A putative Motif I sequence, GLTP, was also identified proximal to the Motif II sequence, immediately preceding the alpha-3 fold. Other HPV types contained a Motif II sequence at the same position as in HPV-16 within their beta-2 sub-domains, e.g. HPV-11 (HIQ), HPV-31 (HLQ) and HPV-33 (HLQ), but contained an arginine (rather than the lysine residue typical of Motif III elements of RCR initiator proteins) within their Motif III-like sequences as follows: HPV-11 (YWFR), HPV-31 (YWYR) and HPV-33 (YWFR). The BPV-1 E1 protein sequences at the corresponding positions in the beta-2 and alpha-5 folds are QMQ and FWYK, and are predicted to be inert with respect to endonuclease potential. These observations are of interest in view of a previous study which showed that HPV-16 switched to a RCR mode in a keratinocyte cell line when the cells were induced to undergo terminal differentiation [Flores & Lambert (1997), J. Virol. 71, 7176-7179]. Evidence for a rolling circle mode of replication for BPV-1 has previously been published (see references below).