Project 1. Characterisation of the functional interaction between HPV16 E2 and Rint-1 AND Project 2. Role of adenovirus E1A in viral gene regulation

Nazeer, Reshma (2014). Project 1. Characterisation of the functional interaction between HPV16 E2 and Rint-1 AND Project 2. Role of adenovirus E1A in viral gene regulation. University of Birmingham. M.Res.

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Project 1:

Human papillomavirus (HPV) infection causes wide range of infections in human beings resulting in 100% of cervical cancers and other anogential and oropharyngeal cancers. HPV protein E2 plays a vital role in subsequent productive and persistent viral infections; HPV E2 protein being a multi functional protein binds to several cellular proteins involved in viral entry pathways and seems to have a role in modulating intracellular trafficking. Previous work in the Parish lab has isolated an interaction between HPV16 E2 and Rint-1 and hypothesized that the association of HPV E2 with membrane associated protein Rint-1 is a part of a novel mechanism in which HPV modulates sub-cellular trafficking during HPV infection. The aim of this project is to further investigate the interaction between Rint-1 and HPV E2 and we successfully characterised the functional interaction between Rint-1 and HPV16 E2 in vivo and from the results obtained, we confirmed that Rint-1 interacts with N-terminus of E2. We also tried to analyse why Rint-1 is interacting with E2 in vivo but not in vitro with phosphatase treated IP. We also determined the localisation of HPV16 E2 protein and Rint-1 protein in sub-cellular elements by sub-cellular fractionation and confirmed that Rint-1 and E2 together localise in cytoskeletal fraction. We also tried to analyse the localisation of HPV16 E2 following siRNA mediated depletion of Rint-1 by Knockdown experiment. From this work, we can speculate that associations of E2 with Rint-1 will have a role in early stages of HPV infection.

Project 2:

Adenoviruses are used as a model system to study the transcription of viral and cellular genes during viral infection, and dissect the molecular basis of cellular transformation. The Adenovirus (Adv) early gene product, 13S E1A serves to transactivate viral early genes; the CR3 domain of E1A plays a vital role in this process. The aim of this project was to investigate further the role of E1A interaction with the cellular histone-directed H3 K4 methyltransferases, Set1A and Set1B, in transactivation. We successfully confirmed by reciprocal co-immunoprecipitation in HEK 293 cells, the in vivo association of Set1A and Set1B with Adv E1A. We also determined by chromatin immunoprecipitation that Set1A and Set1B associate with viral early gene promoters in Adv5-infected cells, suggesting that Set1A and Set1B might be important for transactivation function. We also demonstrated that Adv E1A associates with functional histone H3- and H4- directed methyltransferase activities in HEK 293 cells. We also identified a number of, potentially new, E1A-interacting proteins by mass spectrometry, of which E1A association with MCM proteins was validated in HEK 293 cells. This work has established potential new roles for Set1A and Set1B in E1A transactivation, and, identified a number of novel E1A-interacting proteins.

Type of Work: Thesis (Masters by Research > M.Res.)
Award Type: Masters by Research > M.Res.
College/Faculty: Colleges (2008 onwards) > College of Life & Environmental Sciences
School or Department: School of Biosciences
Funders: None/not applicable
Subjects: Q Science > QR Microbiology > QR355 Virology


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