Dissertaion Defense Announcement
To: The George Mason University Community
Candidate: Beatrix W. Meltzer
Program: PhD Biosciences
Date: Monday April 7, 2014
Time: 10:00 a.m.
Place: George Mason University
Bull Run Hall, Room 130
Title: “Regulation of Preintegration Transcription by Tat and Cellular Transcription factors NF-kappa B and Sp1”
Dr. Yuntao Wu, Chair
Dr. Kylene Kehn-Hall, Dr. Barney Bishop, Dr. Jia Guo Committee Members
For a productive life cycle, HIV-1 needs to integrate its genome into the human cellular chromatin. Studies have shown, however, that prior to integration, the unintegrated viral genome is transcriptionally active. The role of cellular transcription factors, such as Sp1 and NF-ęB, and the viral protein Tat with respect to regulation of preintegration transcription is not yet understood. This early transcriptional activity also generates three regulatory proteins: Tat, Nef, and Rev. Nef from unintegrated viral DNA has been shown to downregulate the cell surface expression of CD4, CXCR4, CCR5, and MHC class I. However, the functional importance of Tat produced from preintegration transcription in the viral life cycle is unclear.
In this dissertation, I demonstrate that deleting either Tat or TAR from the HIV-1 viral genome causes a decrease in the packaging of the virion genomes, resulting in a decrease in the subsequent viral reverse transcription and preintegration transcription in target cells. The Tat defect can be complemented by providing Tat in trans in the virion producer cells. I also demonstrate that the basal transcription from nonintegrated viral DNA is regulated by Sp1 and NF-ęB; mutations in the SP1 and NF-ęB binding sites decrease preintegration transcription, and mutations in all three Sp1 binding sites cause the greatest impairment in preintegration transcription. I further demonstrate that Tat produced from unintegrated viral DNA is functional in stimulating HIV-1 LTR. Nevertheless, the nonintegrated viral DNA is less responsive to Tat transactivation than integrated provirus. I also confirm that nonintegrated viral DNA is assembled into a minichromatin which may inhibit preintegration transcription in the absence of Tat. Thus, the early Tat produced from nonintegrated viral DNA is functionally important in maintaining an open chromatin for persistent low-level transcription, which can modulate cellular activity to promote infection.