Genetic and Genomic Analysis of Tumor Virus Infection and Pathogenesis
My laboratory is interested in the molecular pathogenesis of tumor viruses. We use Kaposi’s sarcoma-associated herpesvirus (KSHV) as a model system to study how an oncogenic virus modulates cellular pathways to facilitate its infection and replication, and cause cancers. KSHV is the etiologic agent of Kaposi’s sarcoma, a vascular spindle angiogenic cancer of proliferating endothelial cells, commonly found in patients with AIDS in Western countries, and in both adult and pediatric patients with cancers in Africa.
Our early efforts were focus on the epidemiology of KSHV when it was a newly identified as a human pathogen. We developed the first-generation serologic assays for the detection of KSHV infection, conducted several important KSHV epidemiologic studies that demonstrated the association of KSHV infection with the development of Kaposi's sarcoma, and defined the epidemiology of KSHV infection in the general populations in the Western countries and Africa. We later extended the studies to South Texas, and showed that KSHV seroprevalence and genotypes in this region are different from other regions in US. We have also developed a genotyping technique that is useful for studying KSHV person-to-person transmission by tracking individual KSHV isolates.
In the last seven years, we have shifted our focus to the molecular basis of KSHV-induced pathogenesis. We initially identified and characterized the first KSHV oncogene viral interferon regulatory factor (vIRF). vIRF is the first viral protein identified to have sequence homology to cellular IRFs, a family of transcriptional factors that are involved in a variety of functions including but not limiting to oncogenesis, cell proliferation, differentiation, apoptosis, and host defense. We have defined the transcriptional mechanism controlling vIRF expression and identified a novel transcriptional silencer, which could have implication for the transcriptional repression of other KSHV lytic genes in KSHV latency.
We identified and characterized the KSHV immunodominant major latent protein, LNA (LANA). LNA has since been shown to be a multi-functional viral protein that maintains the stability of viral episomes, targets tumor suppressor pathways, and regulates viral and cellular gene transcriptions. We have identified a novel nuclear protein KLIP1 that interacts with LNA, and shown that KLIP1 is a cell cycle-dependent potent transcriptional repressor manipulated by LNA in KSHV latent infection. A recent study indicates that KLIP1 interacts with myeloid leukemia factor 1 (MLF1), and possibly has a role in the genesis of erythroleukemias.
The lack of an efficient infection system and a genetic manipulation system has tremendously impeded the advancements of the KSHV field in the last decade. In the last two years, we have developed an efficient bacteria-mammalian shuttle system for KSHV genetic manipulation by cloning the full-length KSHV genome as a bacteria artificial chromosome (BAC). In the same time, we have established a highly efficient KSHV infection system of primary human umbilical vein endothelial cells (HUVEC). The developments of these systems have enabled us to employ a comprehensive genetic, genomic, molecular, cellular, and biochemical approach to define the molecular basis of the sophisticated interplays of virus-cell interactions. To this end, we have performed whole KSHV genome real-time RT-PCR array and cellular microarray analysis and identified essential viral and cellular genes/pathways in KSHV primary infection. We have shown that efficient KSHV infection of HUVEC is productive at the early stage of infection but the virus switches to latency at the later stage of infection. Unsurprisingly, KSHV infection modulates multiple cellular pathways, including MAPK pathways, to facilitate its initial entry and infection of cells. Significantly, KSHV infection suppresses the overall host transcriptional program but selectively activates cellular genes to promote cell growth and cell cycle progression, enhance cell adhesion and invasiveness, and induce inflammatory and angiogenic cytokines. Importantly, we have shown that KSHV infection induces chromosome instability, which could predispose the infected cells to transformation. Our current major efforts are to identify KSHV genes that are essential for malignant cellular transformation. We have constructed a transposon mutagenesis library that contains mutants of all known KSHV genes. The current focus is on three KSHV major latent genes LNA, vFLIP and vCyclin. We have defined the essential function of LNA in episomal persistence, and identified novel functions for vFLIP and vCyclin in the growth and survival of KSHV-infected cells.