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The laboratory of Patrick Green is internationally recognized for their contributions to the understanding of the molecular basis of T-lymphocyte transformation and induction of leukemia/lymphoma and neurological disease by the human T-cell leukemia viruses (HTLVs). The Green lab has three areas of research focus investigating viral and cellular regulators of HTLV gene expression/replication, cellular transformation, and virus survival or persistence in the infected host.
One aim of our laboratory is to understand the mechanism of action and biological role of a unique HTLV-1 accessory gene, termed Hbz. It is the only gene transcribed from the antisense strand of the viral genome and is expressed in almost all adult T-cell leukemia (ATL) cells, whereas tax oncogene expression is typically undetectable. We have discovered that HBZ is dispensable in cell culture, but is required to enhance virus replication and survival in the infected host. More recently utilizing a Hbz shRNA knockdown approach and NOD/SCIDγc-/- (NOG) mice we discovered that Hbz expression enhances the proliferative capacity of HTLV-1 infected cells in culture and plays a critical role in infected cell survival and ultimately HTLV-1 tumorigenesis. HBZ, originally thought to be unique to HTLV-1 has been hypothesized to play a role in pathogenesis. Recently, an antisense protein of HTLV-2 (APH-2) was identified. Despite its lack of a typical b-ZIP domain, APH-2, like HBZ, interacts with CREB and down-regulates Tax-mediated viral transcription. We will investigate in detail the mechanism of action of APH-2. Importantly, since HTLV-1 and HTLV-2 are closely related retroviruses, but have distinct etiological roles in human disease, we hypothesize that comparative studies on anti-sense proteins of HTLV-1 and HTLV-2 will provide fundamental insights into their distinct pathogenic properties.
A second emphasis of our laboratory is to understand the mechanism(s) of post-transcriptional control of viral replication which is critical for HTLV persistence in the infected host. We have initially focused our studies on the HTLV regulatory protein Rex which is a critical on/off switch for viral replication and has been implicated in the transition from early-to-late phase of HTLV gene expression and potentially influences viral latency and long term survival in the infected host. We have discovered and are characterizing a novel carboxy terminal inhibitory domain of Rex that upon phosphorylation positively regulates Rex function. Mutational analysis and proteomic approaches are currently underway to precisely map key phosphorylated amino acid residues and to identify the cellular kinase/phoshatase involved. Furthermore, we have discovered that one of the HTLV accessory proteins, HTLV-1 p30 and the related HTLV-2 p28, functions to repress viral replication by a novel post-transcriptional mechanism. These proteins specifically bind and retain the mRNA of key positive viral regulators in the nucleus leading to reduced protein expression and virion production. Current studies are aimed at identifying host proteins that interact with p30 and p28 in order to understand their role in pathogenesis. Affinity-tag purification coupled with mass spectrometric (MS) analysis has revealed 37 and 18 interacting cellular partners of p30 and p28, respectively. Validation of these proteins is currently underway.
A third focus of our laboratory is to understand the mechanism(s) by which the HTLV Tax oncoprotein induces cellular transformation. These studies emphasize the use of full-length infectious molecular clones of HTLV-1 and the related less pathogenic HTLV-2, primary human T lymphocytes, and a rabbit animal model. We have previously shown that although Tax is the critical oncoprotein, which is indispensable for viral replication and cellular transformation, but it is the viral envelope that dictates the preferential transformation tropism (CD4 vs CD8 T-cells) of HTLV-1 and HTLV-2. Current studies are underway to dissect the envelop domain responsible for this distinct transformation tropism. Preliminary data suggests that this is a post entry event involving selection and clonal outgrowth.