Association of Human Papillomavirus 16 E2 with Rad50-Interacting Protein 1 Enhances Viral DNA Replication

Abstract

Rad50-interacting protein 1 (Rint1) associates with the DNA damage response protein Rad50 during the transition from the S phase to the G2/M phase and functions in radiation-induced G2 checkpoint control. It has also been demonstrated that Rint1 is essential in vesicle trafficking from the Golgi apparatus to the endoplasmic reticulum (ER) through an interaction with Zeste-White 10 (ZW10). We have isolated a novel interaction between Rint1 and the human papillomavirus 16 (HPV16) transcription and replication factor E2. E2 binds to Rint1 within its ZW10 interaction domain, and we show that in the absence of E2, Rint1 is localized to the ER and associates with ZW10. E2 expression results in a disruption of the Rint1-ZW10 interaction and an accumulation of nuclear Rint1, coincident with a significant reduction in vesicle movement from the ER to the Golgi apparatus. Interestingly, nuclear Rint1 and members of the Mre11/Rad50/Nbs1 (MRN) complex were found in distinct E2 nuclear foci, which peaked during mid-S phase, indicating that the recruitment of Rint1 to E2 foci within the nucleus may also result in the recruitment of this DNA damage-sensing protein complex. We show that exogenous Rint1 expression enhances E2-dependent virus replication. Conversely, the overexpression of a truncated Rint1 protein that retains the E2 binding domain but not the Rad50 binding domain acts as a dominant negative inhibitor of E2-dependent HPV replication. Put together, these experiments demonstrate that the interaction between Rint1 and E2 has an important function in HPV replication.IMPORTANCE HPV infections are an important driver of many epithelial cancers, including those within the anogenital and oropharyngeal tracts. The HPV life cycle is tightly regulated and intimately linked to the differentiation of the epithelial cells that it infects. HPV replication factories formed in the nucleus are locations where viral DNA is copied to support virus persistence and amplification of infection. The recruitment of specific cellular protein complexes to these factories aids efficient and controlled viral replication. We have identified a novel HPV-host interaction that functions in the cellular response to DNA damage and cell cycle control. We show that the HPV E2 protein targets Rad50-interacting protein 1 (Rint1) to facilitate virus genome replication. These findings add to our understanding of how HPV replicates and the host cell pathways that are targeted by HPV to support virus replication. Understanding these pathways will allow further research into novel inhibitors of HPV genome replication.

The MRE11–RAD50–NBS1 Complex Conducts the Orchestration of Damage Signaling and Outcomes to Stress in DNA Replication and Repair

“…HPV-31 increases MRN complex half-life for productive viral replication (215). HPV-16 encodes transcription and replication factor E2 that interacts with RAD50-interacting protein 1 (Rint1) (216). Overexpression of exogenous Rint1 enhances HPV-16 replication, and a truncated mutant that lacks a RAD50-interacting domain reduces HPV-16 replication.…”

E3 Ligase ITCH Interacts with the Z Matrix Protein of Lassa and Mopeia Viruses and Is Required for the Release of Infectious Particles

“…It is possible that this protein plays a similar role during MOPV or LASV infection, by promoting internalization of the alpha-dystroglycan, which acts as a reservoir for these viruses, to facilitate viral entry steps or limit the cell infection and subsequent over-stimulation of innate immune responses. The cellular RINT1 protein, which promotes human papillomavirus 16 replication, was shown to only interact with LASV Z protein in our Y2H assay [53]. The fact that this protein is known to mediate vesicle budding from the Golgi apparatus to the endoplasmic reticulum (ER) suggest that protein and membrane trafficking are important host-cell processes which could be possibly affected during arenavirus infection.…”