Integrase-LexA fusion proteins incorporated into human immunodeficiency virus type 1 that contains a catalytically inactive integrase gene are functional to mediate integration
Ml. Holmes-son et Sa. Chow, Integrase-LexA fusion proteins incorporated into human immunodeficiency virus type 1 that contains a catalytically inactive integrase gene are functional to mediate integration, J VIROLOGY, 74(24), 2000, pp. 11548-11556
Purified fusion proteins made up of a retroviral integrase and a sequence-s
pecific DNA-binding protein have been tested in in vitro assays for their a
bility to direct integration into specific target sites. To determine wheth
er these fusion proteins can be incorporated into human immunodeficiency vi
rus type 1 (HIV-1) and are functional to mediate integration, we used an in
trans approach to deliver various integrase-LexA proteins to an integrase-
defective virus containing an integrase mutation at aspartate residue 64. I
ntegrase-LexA, integrase-LexA DNA-binding domain, or N- or C-terminally tru
ncated integrase-LexA proteins were fused to the HIV-1 accessory protein, V
pr. Coexpression of the Vpr fusion proteins and an integrase-defective HIV-
1 molecular clone by a producer cell line resulted in efficient incorporati
on of the fusion protein into the integrase-mutated virus. In addition, eac
h of these viruses was infectious and capable of performing integration, as
determined by two independent cellular assays that measure reporter gene e
xpression. With the exception of the N-terminally truncated integrase fused
to LexA, which was at about 1%, all of the fusion proteins restored integr
ation to a similar level, at 17 to 24% of that of the wild-type virus. The
low level observed with the N-terminally truncated integrase fused to LexA
is consistent with previous results implying that the N terminus of integra
se is involved in multiple steps of the retroviral life cycle. These data i
ndicate that the integrase-fusion proteins retain catalytic function in the
integrase-mutated viruses and demonstrate the feasibility of incorporating
integrase fusion proteins into HIV-1 for the development of site-directed
retroviral vectors.