Genomic cloning and complete sequence analysis of a highly divergent African human immunodeficiency virus isolate.

Analysis of the complete sequence of a human immunodeficiency virus (HIV) isolate (Ant70) obtained from a Cameroonian patient indicates that this virus is the most divergent strain within the HIV-1 family hitherto described. Comparison of the Pol protein, usually highly conserved within the HIV-1 family, shows only about 73% similarity with the HIVmm isolate, whereas for the more variable proteins such as envelope, similarities of 50% or lower are found. The principal neutralizing determinant (V3 loop) and the immunodominant region within gp41 also contain some unusual substitutions, which may have implications for protein function as well as for serological assays based on these regions. Phylogenetic analyses show that this isolate occupies a unique position relative to the human HIV-1 isolates and the recently described SIVcpz virus, indicating that this Cameroonian isolate may provide us with new insights into the origins of the HIV-1 family.

Human T cell clone identifies a potentially protective 54-kDa protein antigen of Toxoplasma gondii cloned and expressed in Escherichia coli.

Protective immunity against Toxoplasma gondii is recognized to be cell mediated and IFN-gamma is considered to be the major mediator of resistance. Thus, protective Ag of the parasite must induce IFN-gamma-producing T cells. In order to identify such Ag, we have constructed a T. gondii cDNA library in the cloning/expression vector lambda gt11, screened this library with a pool of sera of immune donors, and further screened the set of selected recombinant Ag using, as probe, a T. gondii-reactive T-cell clone (TCC) derived from an infected/immune individual and producing a high level of IFN-gamma. One recombinant Ag was shown to induce TCC proliferation and was characterized. The corresponding mature T. gondii Ag has an apparent molecular mass of 54 kDa and the sequence of the cDNA clone suggests that it is membrane associated. The epitope defined by the TCC on this Ag was found to be present in three Toxoplasma strains independently of their phenotype (virulent or cyst forming). Recognition of this Ag by the TCC was shown to be restricted by HLA-DPw4, the most frequent allele in the Caucasian population (approximately 40%). The use of this Ag as a vaccine component is proposed.

The development of specific rRNA-derived oligonucleotide probes for Haemophilus ducreyi, the causative agent of chancroid.

Part of a ribosomal ribonucleic acid (rRNA) cistron of Haemophilus ducreyi was enzymically amplified using conserved primers within the rRNA molecules, cloned in a plasmid vector, and sequenced. From the nucleotide sequence, eight oligonucleotides complementary to different regions in the 16S and 23S rRNA molecules were selected, chemically synthesized, and used as hybridization probes. Hybridization experiments with at least 41 H. ducreyi strains and 13 or 14 non-H. ducreyi strains revealed that all eight oligonucleotide probes were highly reliable and completely specific for H. ducreyi strains. Comparisons of 16S rRNA sequences confirm that H. ducreyi is a member of the Pasteurellaceae though not closely related to other species in this family.

The androgen-dependent rat prostatic binding protein: comparison of the sequences in the 5' part and upstream region of the C1 and C2 genes and analysis of their transcripts.

The complete gene encoding the polypeptide C1 of the complex androgen-controlled prostatic binding protein was isolated from a rat genomic library. A new genomic fragment (C2B) containing only the 5' part of a C2-related gene was also purified. The segments containing exon 1 and a large part of the adjacent sequences were analysed and compared with the corresponding region of the C2A gene which has been completely sequenced previously. The high structural similarity extending over a large part of all three genomic fragments suggests the duplication of a common ancestral gene, followed by a more recent duplication of the C2-coding region. However, since the structural similarity upstream of position -150 between C2A and C2B abruptly disappears and no transcripts specific for the C2B region can be detected in prostate RNA, we propose that at a later stage in evolution the C2B region was disrupted and inactivated. Despite the common origin and the similar regulation of the two active genes, C1 and C2A, the only obvious conserved structural element is the homopurine stretch located at position -400, although sequence motifs resembling steroid hormone response elements are present at several locations.

Rat prostatic binding protein: the complete sequence of the C2 gene and its flanking regions.

The complete sequence (2879 bp) of the androgen-controlled rat prostatic binding protein C2 gene and 1023 bp of the 5'- and 2127 bp of the 3'-flanking regions have been determined. The gene contains three exons (93, 203 and 147 bp) and two introns (1630 and 806 bp). It is flanked by two homopurine-homopyrimidine stretches of 55 and 131 nucleotides respectively, located at positions -405 and 4151. These sequences are remarkably sensitive towards S1-nuclease, indicating an altered DNA conformation under superhelical stress. Several palindromes and dyad structures are observed in the 5'-upstream region of the gene and at position -457, and 80% homology to the consensus sequence of a glucocorticoid receptor binding site is found.