Cellular targets of Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen
The latency-associated nuclear antigen (LANA1) of Kaposi's sarcoma-associated herpesvirus (KSHV) is a multi-function protein involved in maintenance of the viral episome and has been shown to interact with several proteins including p53 and pRB. It is likely that the multifunctional role of LANA1 exceeds these observations therefore the work in this thesis aimed to study LANA1 at both the transcription and protein-protein interaction level. I developed a lentiviral system for the infection of primary endothelial cells with LANA1, to analyse changes in gene expression profiles by gene expression microarrays. While the system was successfully developed, no significant changes in gene expression could be attributed to LANA1. Subsequently, using the yeast two-hybrid system and large-scale immunoaffinity purification coupled to mass spectrometry, I identified 26 novel binding partners of the KSHV LANA1 complex. The majority of these proteins have functions in splicing or mRNA processing and further analysis lead to the discovery of predominant protein domains. Several of the identified proteins, including heterogeneous nuclear ribonucleoprotein (hnRNP) A1, A2/B1, D and I, constitute members of the human H-complex. hnRNP A1, A2/B1 and D are also implicated in telomere biogenesis. I show that LANA1 binds UP1, the proteolytic derivative of hnRNP A1, which modulates telomere elongation and maintenance. Furthermore, I show an in vivo interaction between LANA1 and human telomerase reverse transcriptase (hTERT) and the ability of LANA1 to recover telomerase activity from cell lysates. These findings suggest a function for LANA1 in the maintenance of telomeres and may have important implications for the role of LANA1 in KSHV-related tumours. I propose models for the role of these complexes in splicing and telomere biogenesis.