Codon usage optimization of HIV type 1 subtype C gag, pol, env, and nef genes: in vitro expression and immune responses in DNA-vaccinated mice.
Codon usage optimization of human immunodeficiency virus type 1 (HIV-1) structural genes has been shown to increase protein expression in vitro as well as in the context of DNA vaccines in vivo; however, all optimized genes reported thus far are derived from HIV-1 (group M) subtype B viruses. Here, we report the generation and biological characterization of codon usage-optimized gag, pol, env (gp160, gp140, gp120), and nef genes from a primary (nonrecombinant) HIV-1 subtype C isolate. After transfection into 293T cells, optimized subtype C genes expressed one to two orders of magnitude more protein (as determined by immunoblot densitometry) than the corresponding wild-type constructs. This effect was most pronounced for gp160, gp140, Gag, and Pol (>250-fold), but was also observed for gp120 and Nef (45- and 20-fold, respectively). Optimized gp160- and gp140-derived glycoproteins were processed, incorporated into virus particles, and mediated virus entry when expressed in trans to complement an env-minus HIV-1 provirus. Mice immunized with optimized gp140 DNA developed antibody as well as CD4+ and CD8+ T cell immune responses that were orders of magnitude greater than those of mice immunized with wild-type gp140 DNA. These data confirm and extend previous studies of codon usage optimization of HIV-1 genes to the most prevalent group M subtype. Our panel of matched optimized and wild-type subtype C genes should prove valuable for studies of protein expression and function, the generation of subtype-specific immunological reagents, and the production of DNA-based sub-unit vaccines directed against a broader spectrum of viruses.
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Feng
F
Gao
Yingying
Y
Li
Julie M
JM
Decker
Fred W
FW
Peyerl
Frederic
F
Bibollet-Ruche
Cynthia M
CM
Rodenburg
Yalu
Y
Chen
Denise R
DR
Shaw
Susan
S
Allen
Rosemary
R
Musonda
George M
GM
Shaw
Allan J
AJ
Zajac
Norman
N
Letvin
Beatrice H
BH
Hahn
D016915:AIDS Vaccines / Q000276:immunology*; D000818:Animals; D003062:Codon; D005260:Female; D015752:Genes, env; D015750:Genes, gag; D015754:Genes, nef; D015751:Genes, pol; D015497:HIV-1 / Q000145:classification / Q000235:genetics* / Q000276:immunology; D007371:Interferon-gamma / Q000096:biosynthesis; D051379:Mice; D008807:Mice, Inbred BALB C; D019444:Vaccines, DNA / Q000276:immunology*
D016915:AIDS Vaccines; D003062:Codon; D019444:Vaccines, DNA; D007371:Interferon-gamma