Generation and Characterization of Sequence-Independent SUMOylated Lens Epithelium-Derived Growth Factor (LEDGF) Protein p75 Chimeras.

LEDGF/p75 is a transcriptional coactivator required for efficient lentiviral cDNA integration. LEDGF/p75 has been found to be SUMOylated at four lysine residues, and mutation of these residues to arginine inhibits LEDGF/p75 SUMOylation and impairs the HIV-cofactor activity of this protein. However, we still do not understand how SUMOylation influences the HIV cofactor activity of LEDGF/p75. It has been demonstrated that attachment of SUMO to transcription factors in a non-sequence specific fashion recapitulates the effect of sequence-specific SUMOylation on the activity of these proteins. These results have suggested that is SUMO per se and not the modification of the target protein by SUMO what determines the functional implications of SUMOylation. In this project we will generate and characterize the protein stability of LEDGF/p75 chimeras containing SUMO fused in a sequence-independent manner to the C-terminal region of SUMOylation-deficient LEDGF/p75. If these chimeric proteins are stable, they will be used to generate cell lines stably expressing them