MOLECULAR CLONING AND EXPRESSION OF MACAQUE MONKEY CD4 GENES ; FOR ESTABLISHMENT OF AIDS ANIMAL MODELS.

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Although SIV/macaque monkey systems are known to provide useful animal models for the study of AIDS,SIV resembles HIV-2 in respect of its genetical elements more than HIV-1, of which prevalence is worldwide. Suitable animal models are clearly prerequisite for developements of effective vaccines and therapies against HIV-1, but are unfortunately few. So far SIV and HIV-1 have been extensively studied from the molecular aspects to elucidate the mechanisms determining its host range. In contrast, studies on the cellular receptor (s) of hosts, macaque monkeys are limited until now. We have reported that cynomolgus monkey CD4 could serve as a receptor for HIV-1 when expressed on a human cell line and that another cofactor (s) other than CD4, which exsits in a human cell but not in a monkey cell, might be required for efficient entry of HIV-1 into susceptible host cells. Therefore, it might substantially determine the narrow host range of HIV-1. To address this further, we make an attempt of … More molecular cloning of several macaque monkey CD4s to elucidate the relationship between their V1 domain structures and susceptibilties to HIV-1. We isolated monkey CD4 genes from the thymocyte or PBL libraries of several species of monkeys such as rhesus, pig-tailed, japanese, cynomolgus, african green and squirrel monkeys, which show high homology, depending upon their phylogenic proximity, to the human counterpart through the entire coding region but some critical differences exist in CDR regions of V1 domain, which is thought to constitute essential binding sites for gp120 of HIV-1. As these differences are suspected to reflect the unsusceptibility of monkeys to HIV-1, we constructed an array of mammalian expression vectors containing each monkey CD4 gene, transfected a human cell line HeLa under the seletion pressure of G418 and established several stable transformants expressing each monkey CD4 molecule. These transformants were then exposured with an infectious molecular clone HIV-1NL432 to study whether or not each monkey CD4 can serve as a receptor for the establishment of HIV-1 entry by IFA,syncythium assay and PCR detecting proviral genomic DNA.HeLa expressing monkey CD4 derived from japanese and african green monkeys could not only support the replication of HIV-1 but also form syncytia due to HIV-1. HeLa expressing monkey CD4 from cynomolgus and rhesus monkeys were also revealed to contain the integrated provirus of HIV-1 but not to form syncytia. In contrast HeLa expressing pig-tailed monkey CD4 showed no evidence of virus replication within their cytoplasm. Taken together, these findings indicated that there might exist some flexibility in the CD4 V1 domain structure for the binding with gp120 of HIV-1, and that syncytia formation might be attributed to mechanisms diffrent from the simple virus-cell fusion events. These experimental system might provide an analytical tool to shed some insight into the interaction between monkey CD4 and HIV-1. Less