Universal cellular tropism?
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CD4-Positive T-Lymphocytes↗
Biomedical subjects
J Goudsmit
Publications and source records attributed to J Goudsmit.
Alternative view on AIDS.
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Acquired Immunodeficiency Syndrome↗
Comparison and fine mapping of both high and low neutralizing monoclonal antibodies against the principal neutralization domain of HIV-1.
Monoclonal antibodies raised against viral lysate of HIV-1 (strain LAV-1) and against recombinant gp 160 of HIV-1 (strain HTLV IIIB) which neutralized HIV-1 in a type specific manner were mapped with the aid of peptides (Pepscan analysis). Each of these monoclonal antibodies bound to peptides located on the principal neutralizing domain (PND) of HIV-1. We found that the antigenic sites of the MAbs described in this paper are represented by linear peptides of at least 10 amino acids long. The affinity of the MAbs is high for these peptides and in the same order of magnitude as for native gp 160. The fine mapping of the epitopes may reflect structural features of the PND, for instance which amino acid side chains are exposed and which are buried in the protein. Furthermore the fine mapping of the epitopes explained the HIV type-specific neutralizing activity of the MAbs. Antibodies that bound to the tip of the loop (amino acids QRGPGRAF) have a higher neutralizing activity than antibodies that bound to amino acids towards the N-terminal side of the loop (amino acids KSIRI). Furthermore, MAbs that bound to virtually the same amino acids on the tip of the loop (amino acids IQRGPGRAF and RGPGRAFV) had different neutralizing activities due to different affinities for native gp 160. These data reveal that neutralizing activity not only is determined by the affinity of an antibody to the neutralizing site but also by its fine binding specificities to the V 3 loop of gp 120.
Amino Acid Sequence↗
Natural variants of the HIV-1 long terminal repeat: analysis of promoters with duplicated DNA regulatory motifs.
Sequence variation in the long terminal repeat (LTR) region of HIV-1 was analyzed in viral isolates of 17 infected individuals. Two classes of LTR size variants were found. One HIV-1 variant was detected containing an additional binding site for the transcription factor Sp1. Another LTR size variation was observed in four patients in a region just upstream of the NF-kappa B enhancer. This variation was the result of a duplication of a short DNA sequence (CTG-motif). Cell culture experiments demonstrated that the natural variant with four Sp1 sites had a slightly higher promoter activity and viral replication rate than the isogenic control LTR with three Sp1 sites. No positive effect of the duplicated CTG-motif could be detected. In order to measure small differences in virus production more accurately, equal amounts of a size variant and the wild-type plasmid were cotransfected into T-cells. The virus with four Sp1 sites did outgrow the three Sp1 virus in 35 days of culture and CTG-monomer virus outcompeted the CTG-dimer virus in 42 days. Based on these results we estimate a 5-10% difference in virus production of the LTR variants when compared to that of wild-type.
Base Sequence↗
HIV-1 genomic RNA diversification following sexual and parenteral virus transmission.
Human immunodeficiency virus type 1 (HIV-1) genomic RNA variation was studied in seven presumed donor-recipient pairs directly following sexual (6/7) or parenteral (1/7) transmission. The first RNA-positive serum sample of each recipient and the serum sample of the virus transmitter, identified by epidemiological history and taken within a time bracket of three months of the recipient seroconversion, were analyzed by polymerase chain reaction amplification followed by sequencing of eight cDNA clones of 276 bp, including the V3 coding region. The sequence populations of the recipients were without exception homogeneous, while the sequence populations of the transmitters showed varying degrees of heterogeneity. Nucleotide distance between consensus sequences of unrelated individuals from the Amsterdam population (interpatient variation) averaged 11% (range 7-15%). The largest distance between two clonal sequences of one individual (intrapatient variation) was also 11%. Consensus sequences of five recipients differed by only 0-1% from the consensus sequence of the presumed transmitter, including two pairs of which the transmission was either proven or highly probable. This contrasted with a difference of 10-12% in two pairs, casting doubt on the epidemiological relatedness. Antibody reactivity to a panel of V3 peptides with varying degrees of similarity to the V3 sequences obtained did not augment the discriminatory power of sequence analysis. Results of the sequential sequencing of samples of one transmitter suggest that this was due to an anamnestic antibody response of the transmitter to early variants. From the loss of sequence heterogeneity following transmission and the consensus sequence similarities observed within five transmitter-recipient pairs, we conclude that HIV-1 transmission results in the selection of a limited number of genomes carrying on the infection in the new host, but does not generally lead to a shift in the sequence population as defined by the consensus sequence.
Amino Acid Sequence↗
Natural HIV-1 NEF accelerates virus replication in primary human lymphocytes.
HIV-1 NEF genes were isolated directly from peripheral blood lymphocyte DNA of two HIV-1-infected individuals and cloned into an HXB-2-infectious molecular clone. The effect of NEF on virus production in T-cell lines and primary human lymphocytes was studied. Naturally occurring NEF accelerates virus production in primary human lymphocytes, but not in T-cell lines.
Amino Acid Sequence↗
Assessment of HIV-1 antigenic diversification using serum as sample source.
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Antigenic Variation↗
HIV-1 tropism: truth or consequences?
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Cell Line↗
Characterization of the specificity of the human antibody response to the V3 neutralization domain of HIV-1.
The major neutralization domain of HIV-1, contained in the third variable region (V3) of the external envelope, is highly variable at positions flanking a conserved glycine-proline-glycine sequence. We investigated the relation between V3 sequences of HIV-1 variants circulating in a host and that host's antibody specificity. Multiple V3 sequences were obtained directly, via PCR and subsequent cloning, from serum RNA or cellular DNA from 26 individuals (from 12 around seroconversion). Then, specificity of sera from these individuals to a panel of V3 peptides was determined. The specificity (best recognized peptide) of the early antibody response accurately reflected the virus population circulating around seroconversion in 12/12 individuals and 4/4 HIV-1-infected chimpanzees. A change in serum specificity at later stages of infection was rare: five years after seroconversion, only 3 of 46 individuals had a specificity that differed completely from that in the first year. However, the V3 domain of the virus does change over time, as evidenced by the poor correlation between V3 sequences obtained late in infection and V3 antibody reactivity at the same time point. Thus, in contrast to the accurate antibody response to HIV-1 variants early after infection, generally a specific response to variants emerging at later stages seemed to be absent or of low level. Instead, the early response appeared to be preserved. Finally, we made use of the observed accurate reflection to analyze the variation for the V3 domain of HIV-1 in the Netherlands by probing specificities of early sera from 129 Dutch seroconverting individuals. Specific reactivity to RKSIHIGPGRAFYTTG was found in 36%, to RKSINIGPGRAFYTTG in 12% and to RKSIPIGPGRAFYTTG in 18% of these Dutch sera.
Amino Acid Sequence↗
Ordered appearance of zidovudine resistance mutations during treatment of 18 human immunodeficiency virus-positive subjects.
The order of appearance in the reverse transcriptase gene of four mutations implicated in the development of resistance to zidovudine was investigated by selective polymerase chain reaction. Serial human immunodeficiency virus isolates were studied from 18 initially asymptomatic individuals who had been treated with zidovudine for 2 years. Most subjects had similar patterns. The first mutation occurred transiently at codon 70; its disappearance was paralleled by the appearance of a mutation at codon 215. Subsequently, in some individuals, the mutation at codon 70 reappeared. During the 2 years of treatment, no mutations developed at codon 219 and only one at codon 67, suggesting that most individuals developed only partly resistant virus. This was confirmed by plaque-reduction assay. Six subjects progressed to AIDS within the 2-year study period, confirming that the development of highly resistant isolates is not required for progression in treated individuals. No clear temporal relationship was found between the development of partial resistance and progression.
Acquired Immunodeficiency Syndrome↗
Dehydroepiandrosterone as predictor for progression to AIDS in asymptomatic human immunodeficiency virus-infected men.
The steroid hormone dehydroepiandrosterone (DHEA) has been reported to protect against certain viral infections in animal models and to be a modest inhibitor of human immunodeficiency virus type 1 (HIV-1) infection in vitro. Serum DHEA levels were determined in 41 asymptomatic HIV-1-seropositive subjects, who progressed to AIDS within 5 years after entering a cohort study, in 41 HIV-1-seropositive controls, who remained asymptomatic, and in 41 HIV-1-seronegative controls. At entry, DHEA levels were higher in the seronegative group (median, 13.3 nmol/l) than in either the seropositive nonprogressors (median, 9.2 nmol/l; P = .01) or the progressors (median, 7.2 nmol/l; P less than .001). DHEA levels in the progressors approximately 5 months before the diagnosis of AIDS were lower than the levels in the nonprogressors after the same follow-up (median, 5.6 vs. 8.8 nmol/l; P = .007). DHEA levels less than 7 nmol/l and CD4+ cell counts less than 0.5 x 10(9)/l both proved to be independent predictors for disease progression in HIV-1-infected men.
Acquired Immunodeficiency Syndrome↗
Concordance of human immunodeficiency virus detection by polymerase chain reaction and by serologic assays in a Dutch cohort of seronegative homosexual men.
In three subgroups of a clinically and socially well defined group of Dutch homosexual men, the prevalence of human immunodeficiency virus type 1 (HIV-1) sequences in seronegative blood samples was studied using the polymerase chain reaction (PCR). In 19 seronegative partners of seropositive persons, no HIV-1 sequences were found by PCR in either early (1984/1985) or more recent (1987) samples. In 42 seronegative persons selected by their high risk for HIV-1 infection, none harbored HIV-1 sequences in either early (1985/1986) or late (1989) samples. In 15 people who seroconverted for HIV-1, only 2 samples collected 3 months before seroconversion were PCR-positive. These persons were also HIV antigen-positive at this time. These data suggest that a latent infection greater than 6 months does not occur and that the combination of HIV antibody and HIV antigen tests is appropriate and conclusive in most cases of HIV-1 infection.
Base Sequence↗
Development of HIV-1 group-specific neutralizing antibodies after seroconversion.
OBJECTIVE: To study the induction of group-specific (gs) neutralizing antibodies to HIV-1 after seroconversion. DESIGN AND METHODS: Serum samples taken sequentially from seven Dutch homosexual men and four British haemophiliacs (anonymous sample, therefore sex not known) before and after seroconversion were tested for neutralizing antibodies effective against five diverse HIV-1 strains. Strains of HIV-1 tested included isolates from the United States, Europe and Africa. RESULTS: The gs neutralizing antibody response varied between individuals. Only five of the 11 individuals studied produced detectable neutralizing antibodies to laboratory-adapted HIV-1 strains (for example, IIIB) within 32 weeks of seroconversion. Most individuals initially produced antibodies effective against US/European isolates; the response then generally broadened to include the more diverse strains, i.e., African. CONCLUSIONS: These results suggest that the gs neutralizing target for HIV-1 is poorly immunogenic in vivo and is probably not highly conserved among diverse HIV-1 strains.
HIV Antibodies↗
Modulation of cell surface molecules during HIV-1 infection of H9 cells. An immunoelectron microscopic study.
OBJECTIVE: To study cell surface molecules and HIV-1 proteins on H9 cells 2 days after infection by immunogold electron microscopy, either in single or in double labelling using combinations of host cell-derived molecules and HIV-1 proteins. DESIGN AND METHODS: The presence of host cell antigens CD3, CD4 and human leukocyte antigen-DR (HLA-DR) and HIV-1 antigens gag p15, p17, p24 and env gp41 was evaluated using immunocytochemistry at the light microscopic level. H9 cells 2 days after infection were processed for conventional transmission electron microscopy and cryo-ultramicrotomy. Leukocyte antigens investigated were CD2, CD3, CD4 (two antibodies), CD5, CD8, CD25, CD30, CD63 antigens and HLA-DR; HIV-1-encoded antigens were gag p24, pol reverse transcriptase, and env gp41 and gp120. Double immunogold labelling was performed using reagents with different sized gold particles. For leukocyte markers, the labelling density of the cell membrane was assessed quantitatively on uninfected and infected H9 cells. RESULTS: Infected cells revealed the presence of gag p24, pol, and env gp41 and gp120 antigens on HIV-1 virions. Uninfected H9 cells showed a random distribution of cell surface molecules, including CD4 antigen, along the plasma membrane. The CD63 antigen, a lysosomal membrane glycoprotein, was located mainly in the cytoplasm of uninfected cells. Cells 2 days after infection showed CD4 labelling on sites where virions were budding from or attached to the cell surface and on free virions. Virions also showed labelling by CD3, CD5, CD25, CD30 and CD63 antibodies and anti-HLA-DR. Compared with uninfected cells, a significantly lower density was found on infected cells in labelling for CD4, CD5 and anti-HLA-DR. A significantly higher density on cells 2 days after infection was seen in CD63 labelling. CONCLUSION: During the first phase of infection host cell molecules concentrate on budding structures and newly generated HIV-1 virions. This phenomenon might contribute to the disappearance of these molecules (like the CD4 molecule) from the cell membrane after infection.
Antigens, CD↗
HIV-1 biological phenotype and the development of zidovudine resistance in relation to disease progression in asymptomatic individuals during treatment.
OBJECTIVE: To determine which parameters are associated with clinical progression during zidovudine treatment of asymptomatic HIV-1-infected individuals. METHODS: Twenty-four initially asymptomatic HIV-1-infected individuals were treated with zidovudine and followed until the development of AIDS or for approximately 3 years. HIV-1 phenotype was determined by cocultivation of patient cells with donor lymphocytes, and by a new assay of direct cocultivation with MT-2 cells. Specific mutations in the HIV-1 reverse transcriptase (RT) gene conferring resistance to zidovudine were detected using a selective polymerase chain reaction. RESULTS: Progression to AIDS was more rapid in individuals harbouring syncytium-inducing (SI) viral isolates or showing a conversion from non-syncytium-inducing (NSI) to SI viral isolates. One out of 20 patients who spent a total of 559 months harbouring an NSI phenotype progressed to AIDS, whereas eight out of 12 patients who spent a total of 223 months harbouring an SI phenotype progressed to AIDS (P < 0.001). There was no significant difference between SI and non-SI isolates in the frequency of five mutations causing zidovudine resistance. However, all SI isolates obtained after 2 years of treatment contained mutations in codons 41 and 215 of the RT gene, whereas only five out of 11 (45%) NSI isolates obtained at that time had this combination of mutations. CONCLUSIONS: Conversion to the SI phenotype cannot be prevented by zidovudine treatment. The presence or appearance of an SI virus heralded disease progression in zidovudine-treated individuals. Further research is required to investigate the relationship between virus phenotype and development of zidovudine resistance.
Cytopathogenic Effect, Viral↗
Analysis of human immunodeficiency virus type 1 LTR-LTR junctions in peripheral blood mononuclear cells of infected individuals.
Circularized DNA species containing two long terminal repeat circle junctions were analysed in peripheral blood mononuclear cells of human immunodeficiency virus type 1 (HIV-1)-infected individuals. The circle junction fragments found could be classified into four groups: fragments containing a normal circle junction, fragments with deletions at the circle junction, fragments containing the primer binding site inserted at the circle junction, and fragments containing insertions at the circle junction derived from other regions of the HIV-1 genome.
Base Sequence↗
Minimal requirements for the human immunodeficiency virus type 1 V3 domain to support the syncytium-inducing phenotype: analysis by single amino acid substitution.
The third variable domain (V3) of the human immunodeficiency virus type 1 external envelope contains determinants of cell tropism, cytopathicity, and infectivity and elicits antibodies able to block infectivity in vitro and in vivo. Our study encompassed point-mutational analysis of HXB-2 viruses containing patient-derived V3 regions and expressing a non-syncytium-inducing, low-replicating phenotype in T-cell line SupT1. The mutation within V3 of a serine at position 306 into an also naturally occurring arginine (S to R) required an additional, naturally occurring mutation at position 320 (aspartate to glutamine, D to Q) or 324 (aspartate to asparagine, D to N) for full expression of the syncytium-inducing, high-replicating (SI) phenotype. The naturally occurring mutation of an aspartate into an arginine at position 320 (D to R) was sufficient for production of the SI phenotype. This study proves that introduction of a positively charged amino acid at position 306 or 320, previously shown to be strongly associated with the SI phenotype in field isolates (R.A.M. Fouchier, M. Groenink, N.A. Kootstra, M. Tersmette, H.G. Huisman, F. Miedema, and H. Schuitemaker, J. Virol. 66:3183-3187, 1992), is minimally required for production of SI viruses. In addition, naturally occurring mutations at residue 324 also modulate the virus phenotype.
Amino Acid Sequence↗
Human immunodeficiency virus type 1 clones chimeric for the envelope V3 domain differ in syncytium formation and replication capacity.
Chimeric human immunodeficiency virus type 1 (HIV-1) molecular clones differing only in the envelope V3 region were constructed. The V3 regions were derived from two HIV-1 isolates with a non-syncytium-inducing, non-T-cell-tropic phenotype and from four HIV-1 isolates with a syncytium-inducing, T-cell-tropic phenotype. When assayed in SupT1 cells, the two chimeric viruses with a V3 region derived from the non-syncytium-inducing isolates did not induce syncytia and showed a low level of replication. The four chimeric viruses with a V3 region derived from the syncytium-inducing isolates did induce syncytia and replicated efficiently in SupT1 cells. In A3.01 cells, which do not support syncytium formation, the V3 loop affected replication similarly. Upon prolonged culture in SupT1 cells, the phenotype of a non-syncytium-inducing, low-replicating chimeric HIV-1 converted into a syncytium-inducing, high-replicating phenotype. Mutations within the usually conserved GPGR tip of the loop, which were shown to be responsible for the conversion into the syncytium-inducing, high-replicating phenotype, had occurred. In vitro mutagenesis showed that coupled changes of amino acids at both sides of the tip of the V3 loop were able to convert the viral phenotype from non-syncytium-inducing, low replicating into syncytium inducing, high replicating. Our data show that the V3 loop is involved in both syncytium forming and replicative capacity of HIV-1.
Amino Acid Sequence↗