Sendai virus-based expression of HIV-1 gp120: reinforcement by the V(-) version.
BACKGROUND
We have established a system for recovering Sendai virus (SeV), a nonsegmented negative strand RNA virus, entirely from cDNA at an extremely high rate, and have succeeded in creating a V(-) SeV whose gene expression was greatly enhanced by the deletion of the nonessential V gene. Because of its extreme medical importance, there has been a strong need for the establishment of a better system to express the gp120 envelope glycoprotein of the human immunodeficiency virus type 1 (HIV-1) in sufficient quantity and purity. It also remains to be established to produce gp120 in in vitro natural host cells for HIV-1 such as human primary blood mononuclear cells, macrophages or established T cell lines.


RESULTS
Using the above system, we created recombinant Sendai viruses expressing the gp120 in CV1 cells, a monkey kidney line. The expression level from the standard V(+) version has already reached 2.2/microg per 10(6) infected cells, which was readily purified from the culture fluid with a recovery rate of about 60%, and has so far appeared to be functionally and serologically authentic. The inserted gp120 gene was stably maintained during numerous passages of the recombinant virus. The V(-) version-based expression was even more robust, consistently reaching over 6.0 microg per 10(6) cells, a level that is one of the highest currently attainable for gp120 production in mammalian cells. Furthermore, a broad host range of SeV allowed gp120 production in all the three natural host cells for HIV-1 described above.


CONCLUSIONS
SeV-based expression serves as a novel choice for producing large quantities of HIV-1 gp120 and will greatly facilitate biochemical, biological and immunological studies of this important glycoprotein.
Yu
D
D
Department of Viral Infection, Institute of Medical Science, University of Tokyo, Japan.
Shioda
T
T
Kato
A
A
Hasan
M K
MK
Sakai
Y
Y
Nagai
Y
Y
D004274:DNA, Recombinant; D015870:Gene Expression; D005822:Genetic Vectors; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics; D018108:Respirovirus / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification
D004274:DNA, Recombinant; D015699:HIV Envelope Protein gp120; D011993:Recombinant Fusion Proteins