Analysis Of The First Exon Sequence Characteristics And Function From Tat Gene Of HIV-1 Recombinant Viruses Prevalent In China

Human Immunodeficiency Virus (HIV) is a member of class Retroviridae and genus Lentiviridae. Because of the high error rate of reverse transcription, it is generally found that many nucleotides substitution (transition and transvertion), insertion, deletion, and gene segment repeat or inversion in provirus genome, which gives the notable characteristics of HIV high diversity. One of the major characteristics of the human immunodeficiency virus type 1 (HIV-1) is its unusually high degree of genetic variability. Until now, the strains of HIV-1 from diverse geographic regions have been divided into three groups- group M (major), O (outlier) and N (new or non-M, non-O). The group M isolates comprising nine "pure" subtypes (A-D, F-H, J and K) and fourteen circulating recombinant forms (CRF01-CRF14) are primarily responsible for the global pandemic.The transactivating factor (Tat) of HIV-1 has been shown to be crucial for HIV replication and to be involved in the pathogenesis of AIDS-associated diseases. The central role played by Tat in HIV replication has made it a focus of attention ever since its discovery in 1985. Lentiviral Tat proteins are small arginine-rich RNA-binding proteins encoded by two exons. Amino acids encoded by the first exon are both necessary and functional for TAR RNA binding and transcriptional activation in vivo. Tat is a short viral protein that is essential for the activation and expression of HIV gene. This function, called transactivation, requires the binding of Tat to a nascent leader RNA hairpin (TAR), located at the 5'-end of all HIV typel (HIV-1) mRNAs .Tat has a number of effect on both HIV-infected and uninfected cells and actively contributes to the pathology of AIDS. Tat provides the first example of regulation of viral gene expression through control of transcriptional elongation. In the absence of Tat, initiation from the LTR is efficient but transcription is impaired because the promoter engages poorly processive polymerases that disengage from the DNA template prematurely. Human immunodeficiency virus type 1 (HIV-1) regulatory protein Tat functions as a transcriptional transactivator to increase the production of full-length viral transcripts.The envelope protein variations of env gene have been much studied before, but theTat gene exon varations, are less studied. The Tat gene exon 1 of CRF08-BC has been from subtype C, while Tat region of CRF07-BC is a portion of the Tat gene exon 1 from subtype B\ The CRF07-BC has a recombination breakpoint in the first exon of tat.In this study, the Tat gene exon 1 sequences of the main circulating strains B'/C are presented using nested PCR. Furthermore, the genetic mutations of Tat gene exon 1 are analyzed by sequence analysis and computer soft. By the study of variations of amino acid in the Tat gene exon 1, the variations of amino acid between CRF07-BC and CRF08-BC are mainly in cysteine-rich region and core region. There are no new recombination models in china. The analysis of gene distances and phylogenetic tree of HIV-l B7C recombinant viruses in Tat exon 1 confirm that HIV-l B'/C recombinant viruses are origin from indinan C subtype.Basing on twice surveys of HIV-l molecular epidemiology, we observe that HIV-l CRF-07BC strains become more prevalent than other subtypes strains in china. Comparing with twice surveys of characteristics of CRF-07BC Tat exon 1 gene variation, The 7, 10, 39, 57 site of Tat exon 1 gene have changed. We find the frequency of site 39 and 57 in Tat exon 1 gene increase with the time of epidemic. At the same time, the 10 site of Tat exon 1 gene declines from proportion 56.7 % to 2.5%.To explore the CRF07-BC Tat exon 1 gene mutation and its effect on function with LTR genes, the Tat protein is expressed in eukaryotic expression system which is more close to mammal than escherica coli. Luciferase assays indicate that the Tat protein expressed in vitro has biological activity and can transactivate the down stream reporter gene of HIV-l LTR.The nine TAR motif sequences are analyzed, the result...