Differential budding efficiencies of human T-cell leukemia virus type I (HTLV-I) Gag and Gag-Pro polyproteins from insect and mammalian cells.
In this study, we examined the ability of human T-cell leukemia virus type I (HTLV-I) Gag and Gag-Pro to assemble immature virus-like particles (VLPs) and bud from insect and mammalian cells. Transmission electron microscopy of insect cells infected with a recombinant baculovirus carrying the entire gag gene revealed that Pr53(Gag) is targeted to the plasma membrane, where it extensively accumulates and forms electron-dense evaginations. However, no particles could be detected either inside the cells or in the culture supernatants. With the Gag-Pro-expressing construct, we observed HTLV-I-specific cytoplasmic proteolysis of the Gag precursor, but again no particle released in the culture supernatants. Transmission electron microscopic analysis of insect cells expressing Gag-Pro polyprotein revealed large vacuoles in the cytoplasm and no budding particles at the plasma membrane. In contrast, human immunodeficiency virus type 1 Gag polyprotein expressed in insect cells is able to release VLPs. These data showed that unlike other retroviruses, Pr53(Gag) is unable to be released as immature VLPs from insect cells. To determine whether the block in particle budding and release is due to an intrinsic property of Pr53(Gag) or the absence of essential cellular factors in insect cells, we expressed Gag and Gag-Pro polyproteins in human 293 cells. The results indicate that Pr53(Gag) and p24 capsid are released within particles into the culture supernatants of human 293 cells. We found that the myristylation of the N-terminal glycine residue is essential for Gag release. Altogether, these results strongly suggest that the proper assembly of HTLV-I particles is dependent on mammalian host cell factors.
Bouamr
F
F
INSERM U443, Equipe Rétrovirus et Transfert génique, Bordeaux 2, Bordeaux Cedex, F-33076, France. fbouamr@ms.cc.sunysb.edu
Garnier
L
L
Rayne
F
F
Verna
A
A
Rebeyrotte
N
N
Cerutti
M
M
Mamoun
R Z
RZ
D000818:Animals; D016367:Baculoviridae; D002460:Cell Line; D002462:Cell Membrane / Q000648:ultrastructure / Q000821:virology; D015683:Gene Products, gag / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D015750:Genes, gag; D015497:HIV-1 / Q000235:genetics; D015368:Human T-lymphotropic virus 1 / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D008322:Mammals; D008854:Microscopy, Electron; D011994:Recombinant Proteins / Q000378:metabolism / Q000648:ultrastructure; D018411:Spodoptera; D014162:Transfection
D015683:Gene Products, gag; D011994:Recombinant Proteins