This proposal will analyze the helper functions of HPV for Adeno-Associated Virus (AAV) replication. The parvovirus AAV has a complex life cycle, as a helper-dependent virus, as an autonomous virus in skin, or as an integrated provirus. It is known that adenoviruses, herpesviruses, and human papillomaviruses (HPV) can serve as helpers for AAV. While the adenovirus and herpesvirus helper genes have been studied, HPV helper genes are largely unstudied. We now find that of the HPV early genes E1 has the highest ability to help AAV replication. Our goal in this proposal is to characterize the extent and mechanism of E1 helper function for AAV. The knowledge we gain on E1 helper function for AAV will have dual utility: 1) AAV and HPV interact in skin and their interaction modulates the risk of cervical cancer. Understanding how HPV helps AAV replication, and the mechanism of action, will provide new insight into the cervical carcinogenesis process. 2) Second, rAAV is gaining increasing popularity as a human gene therapy vector. Understanding HPV helper functions for AAV should provide alternative strategies for rAAV production. To analyze HPV helper function for AAV we will:
Aim #1 : Determine if HPV E1 regulates AAV DNA replication, transcription, and virion production.
Aim #2 : Investigate the mechanism of E1 helper function by mapping the """"""""AAV helper domain"""""""" and """"""""Rep78- interaction domain"""""""" within the HPV E1 protein. E1 mutants will be used to map the protein domains. Knowing the helper and Rep78 interaction domains within E1 protein will be the first step in deciphering El's mechanism of helper action.
Aim #3 : Investigate El's mechanism of action by observing cooperation/ complementation of E1 helper function with the known adenovirus helper genes. ? ? ?
|Cao, M; Zhu, H; Bandyopadhyay, S et al. (2012) HPV-16 E1, E2 and E6 each complement the Ad5 helper gene set, increasing rAAV2 and wt AAV2 production. Gene Ther 19:418-24|
|Bandyopadhyay, Sarmistha; Cao, Maohua; Liu, Yong et al. (2010) HPV E1 up-regulates replication-related biochemistries of AAV Rep78. Virology 402:94-101|
|Kang, Bum Yong; You, Hong; Bandyopadhyay, Sarmistha et al. (2009) Cervical cancer isolate PT3, super-permissive for adeno-associated virus replication, over-expresses DNA polymerase delta, PCNA, RFC and RPA. BMC Microbiol 9:79|
|Bandyopadhyay, Sarmistha; Raney, Kevin D; Liu, Yong et al. (2008) AAV-2 Rep78 and HPV-16 E1 interact in vitro, modulating their ATPase activity. Biochemistry 47:845-56|
|You, Hong; Liu, Yong; Prasad, C Krishna et al. (2006) Multiple human papillomavirus genes affect the adeno-associated virus life cycle. Virology 344:532-40|