Rapid size dependent deletion of foreign gene sequences inserted into attenuated HIV-1 upon infection in vivo: implications for vaccine development.
Live attenuated HIV vaccines offer a means to introduce exogenous sequences into the viral genome to target the virus elimination in vivo. Foreign genes inserted into the nef region of HIV-1 NL4-3 were found to be rapidly deleted following virus infection and/or replication, in a size dependent manner, in the human fetal Thymus/Liver implants of severe combined immunodeficient mouse (SCID-hu) model. When the murine heat stable antigen (HSA) of 283 bp was substituted into HIV-1 nef region, the viral loads in vivo were comparable to the negative control nef attenuated HIV-1, and the reporter HSA gene was not deleted upon infection. However, the murine Thy1.2 gene (505 bp) substituted into the nef attenuated HIV-1, upon infection and replication, deleted 441 bp in vitro and 437 bp in vivo, of the inserted Thy1.2 gene. When the enhanced green fluorescence protein (eGFP) gene (720 bp) was substituted for nef, virus replication was aborted in vivo in the Thy/Liv implants, as seen by the background levels of viral loads, comparable to mock infected implants, and the eGFP gene was deleted. When the herpes simplex virus thymidine kinase gene, HSV-TK (1.15 kbp), or HSA gene, was substituted into the viral vpr gene, TK but not HSA gene was deleted, upon infection in vitro. Moreover, NL-TKI reporter virus with both intact nef and vpr genes shows deletion of TK gene both in vitro and in vivo. Excision of foreign genes occurred within the exogenous segments but not in the viral own regions. These results suggest that larger "suicide" genes introduced via HIV-1 can be deleted upon infection. However, smaller size nucleotide sequences or genes (approximately 300 bp) inserted in place of viral nef or vpr gene may be used to target the virus or its components, for attack and elimination in vivo, and thus have implications for the development of live attenuated HIV vaccines.
Brooks
David G
DG
Department of Molecular and Medical Pharmacology, David Geffen School of Medicine and AIDS Institute, University of California, Los Angeles, 90095, USA.
Cohen
Michael D
MD
Jamieson
Beth D
BD
Poon
Betty
B
Kitchen
Scott G
SG
Chow
Samson A
SA
Chen
Irvin S Y
IS
Zack
Jerome A
JA
Koka
Prasad S
PS
D016915:AIDS Vaccines / Q000235:genetics* / Q000276:immunology; D000818:Animals; D015153:Blotting, Western; D051927:CD24 Antigen / Q000235:genetics; D017930:Genes, Reporter; D015754:Genes, nef / Q000235:genetics; D016334:Genes, vpr / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000235:genetics*; D051379:Mice; D016513:Mice, SCID; D017384:Sequence Deletion; D018800:Thy-1 Antigens / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics; D014613:Vaccines, Attenuated / Q000235:genetics / Q000276:immunology; D014779:Virus Replication
D016915:AIDS Vaccines; D051927:CD24 Antigen; C497890:Cd24a protein, mouse; D018800:Thy-1 Antigens; D014613:Vaccines, Attenuated; D049452:Green Fluorescent Proteins; D013937:Thymidine Kinase