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We investigate the molecular and cellular biology of the pandemic lentivirus HIV-1. HIV-1 is the cause of HIV disease, the end stage of which is AIDS. Some known features of HIV-1 replication are diagrammed above. Many aspects of the life cycle remain mysterious. A focus of our research is to decipher how HIV-1 and related lentiviruses interact with host cell proteins, with a long-range goal of enabling new therapeutic strategies and disease models. We are particularly interested in how HIV-1 evades cellular defenses after it enters the cell, transits to the nucleus, and then uses its integrase enzyme to insert (integrate) a DNA copy of its genome into a host cell chromosome. This integration step is permanent, which prevents eradication of HIV-1 and makes lifelong antiviral therapy necessary. Integration is now known to be an excellent therapeutic target. On the other side of the viral life cycle (assembly of new particles), we are using new RNA tracking methods to investigate how, when and where in the cell newly synthesized genomes are incorporated into newly assembling viruses (encapsidation).
One project studies LEDGF/p75, a cellular protein that interacts with HIV-1 integrase. The above figure illustrates the domain structure of LEDGF/p75 and the modular basis for integrase-to-chromatin tethering. See my publications page for primary research reports that resulted in this model and for reviews. N-terminal domains interact with chromatin and a C-terminal integrase binding domain interacts with HIV-1 integrase. The result is a molecular tether. The bottom panels show LEDGF/p75 (red) and integrase (green) co-localizing on chromatin (blue). RNAi knockdown of LEDGF/p75 untethers integrase, which is then found in the cytoplasm (right).
Certain aspects of lentiviral biology, chiefly their ability to carry out the integration step in non-dividing cells, have led to their modification into gene therapy vectors for use in cells that do not divide actively (most cells in the body). We were the first to derive a lentiviral vector from a non-primate lentivirus (Feline Immunodeficiency Virus or FIV) and we continue to construct and use new kinds of HIV-1 and FIV vectors. We are interested in pursuing novel applications within our laboratory and collaboratively at Mayo and elsewhere.
Gene therapy in the eye. One use under study is for nondividing therapeutic targets in the eye, particularly those involved in glaucoma, which is a leading cause of blindness second only to macular degeneration in this country. A glaucoma gene therapy project has resulted in long-term lentiviral vector-delivered transgene expression in human donor eyes as well as in several experimental models. In the figure above the green is the glow of a jellyfish protein (GFP), the gene for which was transferred permanently into cells into the anterior chamber, targeting an eye sub-organ that regulates intraocular pressure. Therapeutic transgenes that decrease intraocular pressure in experimental models have now been developed in the lab. A long term goal is to treat debilitating eye diseases with gene therapy.
Reagents and viral constructs developed on the lentiviral vectorology side of the lab are often used as tools for investigating basic virology and vice versa. For example, we are interested in adapting our integrase work toward targeting lentiviral vectors in the genome to reduce potential for geno-toxicity in human gene therapy.
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