Ritsuko Inubushi

HIV-1 capsid mutants inhibit the replication of wild-type virus at both early and late infection phases

In‐frame mutations were introduced into various portions of the human immunodeficiency virus type 1 (HIV‐1) gag gene, and potentials of the mutants to suppress the replication of wild‐type HIV‐1 were monitored. In contrast to results obtained with matrix and nucleocapsid mutants, almost all capsid mutants blocked HIV‐1 replication completely in single‐round replication assays. A capsid mutant designated C6b was demonstrated to be one of the most efficient inhibitors for HIV‐1 reported to date, and to be effective at both early and late viral replication phases. T‐cells, which are engineered to express the C6b Gag in response to HIV‐1 infection, were perfectly resistant to HIV‐1.

Inhibition of HIV/SIV replication by dominant negative Gag mutants.

There are several major strategies against HIV/AIDS. Of these, the gene therapy is a novel, challenging, and promising one. The target genes, which have been extensively studied for the potential gene therapy of HIV/AIDS, include those of cellular and viral origins. Especially, trans-dominant negative Tat, Rev, Env, Pol, and Gag mutants of HIV have currently attracted considerable attention. In this brief review, we summarize the nature of the HIV/SIV mutants of this category and discuss their future use for gene therapy with special reference to the dominant negative Gag mutants of HIV-1.

Mapping the genetic determinants of human immunodeficiency virus type 2 for cell tropism and replication efficiency

SummaryTwo distinct infectious molecular clones of human immunodeficiency type 2 (HIV-2) were analyzed for their biological properties in six cell lines. Fourteen chimeric and ten mutant viruses were constructed from these two viral genomes to localize the genetic determinants responsible for the phenotypes. Growth property of the viruses in the cell lines, together with the biochemical data, showed that a major determinant for the viral tropism resides in the env gene. In addition, in some cell lines, the accessory genes vif and nef affected the efficiency of virus replication. Thus, like HIV-1, mutations in the auxiliary and env genes of HIV-2 contributed much to the differences in virological characteristics.

Cleavage of Gag precursor is required for early replication phase of HIV-1.

A mutant of human immunodeficiency virus type 1 (HIV‐1), which is deficient for Gag precursor cleavage and non‐infectious, was characterized with respect to its defective step in the viral replication phase. Upon transfection, the mutant produced a normal level of progeny virions as monitored by electron microscopy and RNA hybridization. Single‐round replication assay demonstrated, in contrast, that the mutant was defective at the early phase of the replication cycle. Furthermore, no viral DNA was detected in the cells infected with the mutant. Taken together, it is concluded that maturation of Gag precursor protein of HIV‐1 is required for an early event(s) before or during a coupled process of uncoating/reverse transcription.

Gag-Pol region determines the tropism of SIVagm for human cells.

Simian immunodeficiency virus isolated from African green monkeys (SIVagm) does not grow in many of human cell lines such as CEM×174, H9, and MT-4, but could replicate in some human cell lines. Sequence of SIVagm responsible for its narrow host range was determined by making and monitoring growth potential of chimeric clones between SIVagm and human immunodeficiency virus type 1 (HIV-1). The results obtained indicated that the gag–pol region determines the observed narrow host range. By monitoring virus DNA synthesis and progeny virion production, the defect(s) of SIVagm in the replication in the restricted cells was demonstrated to be located at early phase.

Cell-dependent replication potentials of HIV-1 gag mutants

An infectious molecular clone of human immunodeficiency virus type 1 (HIV-1), designated pNLaiKH, which is tropic for both lymphocytic and monocytic cells, was constructed. To study the early function of HIV-1 Gag proteins in two types of cells, the mutations known to give host cell-dependent early defects were introduced into pNLaiKH, and the replication potentials and defective replication sites in the cells of the resultant mutants were monitored. All mutants grew in some lymphocytic cells, but not at all in monocytic cells. A nucleocapsid mutant was found to be defective at an early replication phase in all the cell lines to various extent, as expected. In contrast, a matrix mutant and a capsid mutant displayed a replication defect in a producer-cell-dependent manner. These results demonstrated that complex interactions of cell factors and Gag proteins are involved in an early process of HIV-1 replication.

Complete Inhibition of SIVmac Replication by its Capsid Mutants

Mutations were introduced into a genomic region encoding the C-terminal portion of Gag capsid protein of pathogenic simian immunodeficiency virus (SIVmac239). All the mutants generated were defective for virion production and were non-infectious for monkey cells. They all efficiently suppressed the replication of wild type SIVmac in monkey cells. These results were in good agreement with those obtained for human immunodeficiency virus type 1, showing the importance of SIV/monkey model system for studies on Gag.