Virological and cellular roles of the transcriptional coactivator LEDGF/p75

Abstract

The chromatin-associated cellular proteins LEDGF/p75 and LEDGF/p52 have been implicated in transcriptional regulation, cell survival and autoimmunity. LEDGF/p75 also appears to act as a chromatin-docking factor or receptor for HIV-1 and other lentiviruses and to play a role in leukemogenesis. For both the viral and cellular roles of this protein, a key feature is its ability to act as a molecular adaptor and tether proteins to the chromatin fiber. This chapter reviews the emerging roles of LEDGF/p75 and LEDGF/p52 in diverse cellular processes and disease states.

Figure 1

Domain structure and binding partners of LEDGF/p75 and LEDGF/p52.

Figure 2. Structure of NUP 98-LEDGF/p75 fusion proteins

NUP98 contains eight N- terminal GLFG repeats that govern localization of the protein to nuclear GLFG bodies. GLEBS is a sequence within the GLFG repeats that serves as docking site to GLE_2p, the yeast ortholog of human RAE₁ a protein involved in mRNA nuclear export. Nucleoporin RNA binding motif (NRM) is an octapeptide with partial homology to the ribonucleoprotein motif. In the NUP98-LEDGF/p75 fusion proteins (I–III) described in leukemic patients, the NUP98 GLFG repeats and the GLEBS element fuse to LEDGF/p75 segments that contain the IBD.

Figure 3. Co-crystal structure of the HIV-1 IN CCD-IBD interface

The figure was constructed with MacPyMOL from Protein Data Bank file 2BJ4 (www.pdb.org, ref. [13]. LEDGF/p75 contributes most of the amino acid side chains that make direct contact with IN. D366 engages in a pair of essential hydrogen bonds with the main chain amides of E170 and H171 in the alpha-4/5 connector of one IN monomer. Hydrophobic interactions predominate in interactions with the other IN monomer: IN residues W132, W131, A128, L102 form a pocket in the vicinity of IBD residues F406 and V408; this pocket buries IBD residue I365). IN catalytic center resides D64, D116 and E152, the mutation of which produce purely catalytic defects (reviewed in [25]) are shown in the lower half of the figure for each IN monomer. Interaction between the HIV-1 IN NTD and the IBD was previously established by biochemical evidence [53] and Hare et al. have recently solved a co-crystal structure for the LEDGF/p75 IBD complexed with a two-domain fragment of HIV-2 IN (NTD+CCD) [38]. Extensive structural contacts between the IBD and the NTD were identified and characterized [38]. Charged interactions are dominant, with conserved acidic residues in the HIV-2 IN NTD (E6, E10, E13) engaging complementary basic residues in the IBD (K401, R404, R405). Moreover, the NTDs of other lentiviral INs contain the same or closely adjacent glutamic acid residues. Enhancement by LEDGF/p75 of concerted strand transfer activity in vitro was also shown to be impaired by charge-reversing mutation of the basic IBD residues to glutamates in the equine lentivirus (EIAV) IN [38]. This activity could then be partially restored by reciprocal mutation of the acidic IN residues to lysines, verifying that these charged interactions are the main structural feature. See ref. [17] for a recent review of HIV-1 IN structural biology.

Figure 4. Alignment of the core catalytic domains of different lentiviral IN proteins

Alignment of the central part of the CCD for IN proteins from the three lentiviral subgenera (primate, feline, ungulate). Identity is indicated by = and residues with conserved biochemical features by dots. IN alpha helices are indicated below and the segments primarily involved in forming the IBD binding pocket are indicated by bold-face font and brackets. The relative lack of sequence conservation in these regions, e.g., the alpha 4/5 connector, is evident. Figure 2 shows the placement of these protein elements for the interface of HIV-1 IN with LEDGF/p75; catalytic center residues (heavy black arrows here) do not contact the IBD. Although its exact oligomerization state in the PIC is not conclusively established, the weight of evidence is in favor of the enzyme acting as a multimer, with a tetramer likely [4, 15, 30, 31, 37, 39, 40, 46, 84, 96]. In vitro, an IN dimer enables 3′ end processing but a tetramer appears needed for DNA strand transfer activity [30, 37, 46]. Of note, higher order multimers were defined in the Hare et al. co- crystal structure of the IBD with HIV-2 IN_NTD+CCD [38]. Their relevance to the oligmeric state in the virus remains to be determined.