Differences in cytopathogenicity and host cell range among infectious molecular clones of human immunodeficiency virus type 1 simultaneously isolated from an individual.

A cytopathic human immunodeficiency virus type 1 (HIV-1) isolate containing multiple virus genotypes was molecularly cloned, and the biological activity of six randomly selected clones was assessed by transfection into human lymphoid or glial cell lines. Five infectious clones of HIV-1, termed N1T-A through -E, were isolated in this manner. Clones N1T-A, -B, -C, and -E could be distinguished by restriction endonuclease mapping whereas clones N1T-B and -D had identical maps with the enzymes used. Each clone exhibited a distinct host cell range as well as markedly different infection kinetics and cytopathogenic properties when tested in human cell lines of T-lymphocytic, monocytic, and astrocytic origin. In particular, infection with HIV-1 clone N1T-E was characterized by slow kinetics and lack of significant cytopathic effects in acutely and chronically infected cells. Clone N1T-A, similar to the parental isolate N1T, exhibited a wide host cell range, fast kinetics of infection, and high cytopathogenicity. These data indicate that HIV-infected individuals may carry multiple HIV-1 genotypes with distinct cytopathogenic potential and cell tropism. Analysis of virus isolates must take into account the contribution, or masking, of individual virus clones.     

Recombinational analysis of a natural noncytopathic human immunodeficiency virus type 1 (HIV-1) isolate: role of the vif gene in HIV-1 infection kinetics and cytopathicity.

Two molecularly cloned coisolates of human immunodeficiency virus type 1 (HIV-1) have been found to exhibit different phenotypes of viral expression, either rapid and cytopathic (N1T-A virus) or delayed and noncytopathic (N1T-E virus [X. Ma, K. Sakai, F. Sinangil, E. Golub, and D. J. Volsky, Virology 176:184-194, 1990]). To identify the viral genetic elements responsible for these phenotypes, we prepared reciprocal recombinants in different regions of N1T-A and N1T-E viral genomes. Infectivity experiments with the recombinant viruses revealed that the rapid/cytopathic (N1T-A-like) phenotype assorted cleanly with the V1f-coding region and Vif expression. The smallest HIV-1 DNA region that conferred the complete phenotypic switch was a 284-bp NdeI-StuI fragment within the vif open reading frame. Nucleotide sequence analysis revealed a 35-bp deletion starting at nucleotide 218 in the N1T-E vif gene. A 23-kDa Vif protein was detected by immunoblotting using Vif-specific antiserum in extracts of cells infected with N1T-A but not N1T-E virus. No detectable vif protein was found in association with sedimented particles of either virus. Cotransfection of a eucaryotic vif expression plasmid with N1T-E DNA complemented the N1T-E defect; rapid/cytopathic infection similar to that in N1T-A-transfected cells was observed. We conclude that Vif controls the rate, and consequently the cytopathic outcome, of HIV-1 infection.     

Generation of an infectious clone of VR-2332, a highly virulent North American-type isolate of porcine reproductive and respiratory syndrome virus

A full-length cDNA clone of the prototypical North American porcine reproductive and respiratory syndrome virus (PRRSV) isolate VR-2332 was assembled in the plasmid vector pOK(12). To rescue infectious virus, capped RNA was transcribed in vitro from the pOK(12) clone and transfected into BHK-21C cells. The supernatant from transfected monolayers were serially passaged on Marc-145 cells and porcine pulmonary alveolar macrophages. Infectious PRRSV was recovered on Marc-145 cells as well as porcine pulmonary macrophages; thus, the cloned virus exhibited the same cell tropism as the parental VR-2332 strain. However, the cloned virus was clearly distinguishable from the parental VR-2332 strain by an engineered marker, a BstZ17I restriction site. The full-length cDNA clone had 11 nucleotide changes, 2 of which affected coding, compared to the parental VR-2332 strain. Additionally, the transcribed RNA had an extra G at the 5' end. To examine whether these changes influenced viral replication, we examined the growth kinetics of the cloned virus in vitro. In Marc-145 cells, the growth kinetics of the cloned virus reflected those of the parental isolate, even though the titers of the cloned virus were consistently slightly lower. In experimentally infected 5.5-week-old pigs, the cloned virus produced blue discoloration of the ears, a classical clinical symptom of PRRSV. Also, the seroconversion kinetics of pigs infected with the cloned virus and VR-2332 were very similar. Hence, virus derived from the full-length cDNA clone appeared to recapitulate the biological properties of the highly virulent parental VR-2332 strain. This is the first report of an infectious cDNA clone based on American-type PRRSV. The availability of this cDNA clone will allow examination of the molecular mechanisms behind PRRSV virulence and attenuation, which might in turn allow the production of second-generation, genetically engineered PRRSV vaccines.     

Demonstration of two distinct cytopathic effects with syncytium formation-defective human immunodeficiency virus type 1 mutants.

The mechanism of human immunodeficiency virus type 1 (HIV-1) cytopathicity is poorly understood and might involve formation of multinucleated giant cells (syncytia), single-cell lysis, or both. In order to determine the contributions of the fusion domain to syncytium formation, single-cell lysis, and viral infectivity and to clarify the molecular details of these events, insertion mutations were made in the portion of env encoding this sequence in the functional HIV-1 proviral clone HXB2. Viruses produced from these mutant clones were found to have a partial (F3) or complete (F6) loss of syncytium-forming ability in acutely infected CEM, Sup T1, and MT4 T-cell lines. During the early stage of acute infection by F6 virus, there was a loss of the syncytial cytopathic effect, which resulted in increased cell viability, and a 1.9- to 2.6-fold increase in virus yield in the cell lines tested. In the late stage of acute infection, the single-cell cytopathic effect of F6 virus was similar to that of the parental HXB2 virus. The F3 and F6 viruses were also found to have a 1.7- to 43-fold reduction in infectivity compared with the HXB2 virus. The mutant F3 and F6 and parental HXB2 envelope proteins were expressed in vaccinia virus, and the mutant envelope proteins were observed to be defective in their ability to form syncytia. BSC-40 cells infected with vaccinia virus recombinants revealed no differences in kinetics of cleavage, cell surface expression, or CD4 binding capacity of the mutant and parental envelope proteins. These results demonstrate that a loss of syncytium formation results in an attenuation of infectivity and a loss of the syncytial cytopathic effect without a loss of single-cell lysis. These mutants may reflect in tissue culture the changes observed in the HIV isolates in vivo during disease progression, which exhibit marked differences in syncytium production.     

Cloning and characterization of human immunodeficiency virus type 1 variants diminished in the ability to induce syncytium-independent cytolysis.

The phenomenon of interference was exploited to isolate low-abundance noncytopathic human immunodeficiency virus type 1 (HIV-1) variants from a primary HIV-1 isolate from an asymptomatic HIV-1-seropositive hemophiliac. Successive rounds of virus infection of a cytolysis-susceptible CD4+ cell line and isolation of surviving cells resulted in selective amplification of an HIV-1 variant reduced in the ability to induce cytolysis. The presence of a PvuII polymorphism facilitated subsequent amplification and cloning of cytopathic and noncytopathic HIV-1 variants from the primary isolate. Cloned virus stocks from cytopathic and noncytopathic variants exhibited similar replication kinetics, infectivity, and syncytium induction in susceptible host cells. The noncytopathic HIV-1 variant was unable, however, to induce single-cell killing in susceptible host cells. Construction of viral hybrids in which regions of cytopathic and noncytopathic variants were exchanged indicated that determinants for the noncytopathic phenotype map to the envelope glycoprotein. Sequence analysis of the envelope coding regions indicated the absence of two highly conserved N-linked glycosylation sites in the noncytopathic HIV-1 variant, which accompanied differences in processing of precursor gp160 envelope glycoprotein. These results demonstrate that determinants for syncytium-independent single-cell killing are located within the envelope glycoprotein and suggest that single-cell killing is profoundly influenced by alterations in envelope sequence which affect posttranslational processing of HIV-1 envelope glycoprotein within the infected cell.     

In Vivo Evolution of a Novel, Syncytium-Inducing and Cytopathic Feline Leukemia Virus Variant

Studies of feline leukemia virus (FeLV) have illustrated the importance of the genotype of the infecting virus in determining disease outcome. In FeLV infections, as in other retroviral infections, it is less clear how virus variants that evolve from the transmitted virus affect pathogenesis. We previously reported an analysis of the genotypic changes that occur in the viral envelope gene (env) in cats infected with a prototype transmissible FeLV clone, 61E (J. Rohn, M. Linenberger, E. Hoover, and J. Overbaugh, J. Virol. 68:2458–2467, 1994). In one cat, each variant (81T) had evolved, in addition to scattered amino acid changes, a four-amino-acid insertion with respect to 61E. This insertion was located at the same site in the extracellular envelope glycoprotein where the immunodeficiency-inducing molecular clone 61C possesses a six-amino-acid insertion critical for its pathogenic phenotype, although the sequences of the insertions were distinct. To determine whether acquisition of the four-amino-acid insertion was associated with a change in the replication or cytopathic properties of the virus, we constructed chimeras encoding 81T env genes in a 61E background. One representative chimeric virus, EET(TE)-109, was highly cytopathic despite the fact that it replicated with delayed kinetics in the feline T-cell line 3201 compared to the parental 61E virus. The phenotype of this virus was also novel compared to other FeLVs, including both the parental virus 61E and the immunodeficiency-inducing variant 61C, because infection of T cells was associated with syncytium formation. Moreover, in single-cycle infection assays, the 81T-109 envelope demonstrated receptor usage properties distinct from those of both 61E and 61C envelope. Thus, these studies demonstrate the evolution of a novel T-cell cytopathic and syncytium-inducing FeLV in the host. The 81T virus will be valuable for dissecting the mechanism of T-cell killing by cytopathic variants in the FeLV model.     

Cytopathic variants of an attenuated isolate of human immunodeficiency virus type 2 exhibit increased affinity for CD4.

Naturally occurring isolates of human immunodeficiency virus (HIV) have been described which are deficient in their ability to fuse with and kill CD4+ target cells. Although the molecular basis for their attenuation has not yet been defined, several lines of evidence point toward the viral envelope gene as a key determinant of viral pathogenicity. In the present article, we report the biological characterization of two highly cytopathic variants derived by repeated cell-free passage of an attenuated isolate of HIV type 2 (HIV-2), termed HIV-2/ST. Unlike the parental virus, the cytopathic variants were found to infect Sup-T1 cells with great efficiency and to induce both cell fusion and profound killing in these cultures. To determine whether changes in the viral envelope gene were responsible for the observed phenotypic differences, we examined the CD4 binding affinity of these viruses using a novel assay designed to quantitate the binding of fluoresceinated CD4 to viral envelope in its native configuration on the cell surface. The results demonstrated that the affinity of parental HIV-2/ST envelope for CD4 was 2 orders of magnitude reduced, while the cytopathic variants exhibited a high CD4 binding affinity, comparable to that of cytopathic HIV-1 and HIV-2 isolates. From these data, we conclude that the cytopathic potential of HIV depends, at least in part, on its receptor-binding affinity. In addition, our study documents strong selection pressures for viruses with increased CD4 affinity during propagation in immortalized T-cell lines, thus emphasizing the need to study HIV envelope biology in natural target cells.     

Characteristics of a pathogenic molecular clone of an end-stage serum-derived variant of simian immunodeficiency virus (SIV(F359))

End-stage simian immunodeficiency virus (SIV) isolates are suggested to be the most fit of the evolved virulent variants that precipitate the progression to AIDS. To determine if there were common characteristics of end-stage variants which emerge from accelerated cases of AIDS, a molecular clone was derived directly from serum following in vivo selection of a highly virulent SIV isolate obtained by serial end-stage passage in rhesus monkeys (Macaca mulatta). This dominant variant caused a marked cytopathic effect and replicated to very high levels in activated but not resting peripheral blood lymphocytes. Furthermore, although this clone infected but did not replicate to detectable levels in rhesus monocyte-derived macrophages, these cells were able to transmit infection to autologous T cells upon contact. Interestingly, although at low doses this end-stage variant did not use any of the known coreceptors except CCR5, it was able to infect and replicate in human peripheral blood mononuclear cells homozygous for the Delta 32 deletion of CCR5, suggesting the use of a novel coreceptor. It represents the first pathogenic molecular clone of SIV derived from viral RNA in serum and provides evidence that not only the genetic but also the biological characteristics acquired by highly fit late-stage disease variants may be distinct in different hosts.     

Molecular characterization of an attenuated human immunodeficiency virus type 2 isolate.

Naturally occurring strains of human immunodeficiency virus (HIV) can vary considerably in their in vitro biological properties, and such differences may also be reflected in their in vivo pathogenesis. In an attempt to define genetic determinants of viral pathogenicity, we have molecularly cloned, sequenced, and characterized an attenuated isolate of HIV type 2 (HIV-2/ST) that differs from prototype HIV-2 strains in its inability to fuse with and kill susceptible CD4-bearing target cells. A proviral clone, termed JSP4-27, was identified to be transfection competent and to fully exhibit the noncytopathic and nonfusogenic properties of its parental isolate. Nucleotide sequence analysis of this clone revealed a genomic organization very similar to that of cytopathic HIV-2 strains and an overall nucleotide sequence homology of 88 to 90%. Amino acid sequence comparison confirmed the integrity of all major viral gene products in JSP4-27 but identified two amino acid sequence substitutions in its envelope fusion region. To investigate whether these mutations were responsible for the nonfusogenic phenotype of JSP4-27, we amplified, cloned, and sequenced the envelope fusion regions of four additional HIV-2/ST strains, two of which represented in vitro-generated, fusogenic and cytopathic variants of HIV-2/ST. The analysis showed that all HIV-2/ST strains examined, including the fusogenic variants, contained the same amino acid sequence changes. On the basis of these findings, we conclude that the attenuated phenotype of JSP4-27, and that of its parental virus, is not due to a direct alteration of the envelope fusion domain. Our results also show, for the first time, that individual replication-competent proviral clones can be representative of attenuated strains of HIV.     

Biological, molecular, and structural analysis of a cytopathic variant from a molecularly cloned simian immunodeficiency virus.

Some isolates of simian immunodeficiency virus (SIV) have been shown to infect Sup-T1 cells with slow kinetics and in the absence of cytopathic effects, including cell fusion or CD4 down-modulation (J. A. Hoxie, B. S. Haggarty, S. Bonser, J. Rackowski, H. Shan, and P. Kanki, J. Virol. 62:2557-2568, 1988). In the present study, we describe the isolation and characterization of a SIVmac variant, derived from the BK28 infectious molecular clone, that became highly cytopathic for Sup-T1 cells. This variant, termed CP-MAC, exhibited a number of differences from BK28, including (i) an altered tropism which largely restricted its host range to Sup-T1 cells, (ii) the ability to induce cell fusion and CD4 down-modulation, and (iii) a highly stable interaction of its external (SU) and transmembrane (TM) envelope glycoproteins. In addition, a marked increase in the level of surface envelope glycoproteins was observed both on CP-MAC-infected cells and on virions. The CP-MAC env gene was PCR amplified from infected cells, and sequence analysis identified five amino acid changes in SU and six in TM compared with BK28. The introduction of these changes into BK28 was shown to fully reconstitute the biological and morphological properties of CP-MAC. The limited number of mutations in CP-MAC should enable the molecular determinants to be more precisely defined and help to identify the underlying mechanisms responsible for the striking biological and structural alterations exhibited by this virus.     

Molecular basis of non-virulence of Trypanosoma cruzi clone CL-14

We investigated the properties of metacyclic trypomastigotes of non-virulent Trypanosoma cruzi clone CL-14, as compared to the parental isolate CL. In contrast to the CL isolate, which produces high parasitemias in mice, metacyclic forms of clone CL-14 failed to produce patent infection. In vitro, the number of clone CL-14 parasites that entered epithelial HeLa cells, after 1 h incubation, was approximately four-fold lower than that of the CL isolate and at 72 h post-infection intracellular replication was not apparent whereas cells infected with the CL isolate contained large number of parasites replicating as amastigotes. CL isolate metacyclic forms were long and slender, with the kinetoplast localised closer to the nucleus than to the posterior end, whereas clone CL-14 parasites were shorter, with the kinetoplast very close to the posterior end. Cysteine proteinase cruzipain and trans-sialidase activities were lower in CL isolate than in clone CL-14. The surface profile was similar, except that the expression of gp82, the stage-specific glycoprotein that promotes CL isolate mucosal infection in vivo and host cell invasion in vitro, was greatly reduced on the surface of clone CL-14 metacyclic forms. Genistein, a specific inhibitor of protein tyrosine kinase, which is activated in CL isolate by binding of gp82 to its host cell receptor, did not affect host cell entry of clone CL-14. In contrast with CL isolate, the infectivity of clone CL-14 was not affected by phospholipase C inhibitor U73122 but was diminished by a combination of ionomycin plus NH(4)Cl, which releases Ca(2+) from acidic vacuoles. Internalisation of clone CL-14, but not of CL isolate, was significantly increased by treating parasites with neuraminidase, which removes sialic acid from the mucin-like surface molecule gp35/50. Taken together, our data suggest an association between the non-virulence of clone CL-14 metacyclic forms and the reduced expression of gp82, which precludes the activation of signal transduction pathways leading to effective host cell invasion.     

Engagement of CD28 modulates CXC chemokine receptor 4 surface expression in both resting and CD3-stimulated CD4+ T cells

Optimal CD4+ T cell activation requires the cooperation of multiple signaling pathways coupled to the TCR-CD3 complex and to the CD28 costimulatory molecule. In this study, we have investigated the expression of surface CXC chemokine receptor 4 (CXCR4) in enriched populations of CD4+ T PBL, stimulated with anti-CD3 and anti-CD28 mAbs, immobilized on plastic. Anti-CD3 alone induced a progressive down-regulation of surface CXCR4, accompanied by a significant decline in the entry of the HXB2 T cell line-tropic (X4-tropic) HIV-1 clone in CD4+ T cells. Of note, this effect was strictly dependent on the presence in culture of CD14+ monocytes. On the other hand, anti-CD28 alone induced a small but reproducible increase in the expression of surface CXCR4 as well as in the entry of HXB2 HIV-1 clone in resting CD4+ T cells. When the two mAbs were used in combination, anti-CD28 potently synergized with anti-CD3 in inducing the expression of CD69 activation marker and stimulating the proliferation of CD4+ T cells. On the other hand, anti-CD28 counteracted the CXCR4 down-modulation induced by anti-CD3. The latter effect was particularly evident when anti-CD28 was associated to suboptimal concentrations of anti-CD3. Because CXCR4 is the major coreceptor for the highly cytopathic X4-tropic HIV-1 strains, which preferentially replicate in proliferating CD4+ T cells, the ability of anti-CD28 to up-regulate the surface expression of CXCR4 in both resting and activated CD4+ T cells provides one relevant mechanism for the progression of HIV-1 disease.     

The cytopathicity of a simian immunodeficiency virus Mne variant is determined by mutations in Gag and Env.

Previous studies suggested that the rapidly replicating, highly cytopathic, syncytium-inducing (rapid-high/SI) phenotype of simian immunodeficiency virus Mne variants that evolved in macaques inoculated with a slowly replicating, minimally cytopathic, non-syncytium-inducing (slow-low/NSI) molecular clone was not solely the result of changes in the envelope surface protein (Env SU). To define the viral determinants responsible for the change in phenotype, we molecularly cloned a rapid-high/SI variant (designated SIVMne170) derived from the peripheral blood mononuclear cells (PBMCs) of a pig-tailed macaque that was inoculated with a slow-low/NSI molecular clone, SIVMneCL8. SIVMne170 was SI and replicated with faster kinetics and was more cytopathic than the parent SIVMneCL8 in CEMx174 cells. Additionally, SIVMne170 was more cytopathic for the CD4+ T-cell population than SIVMneCL8 in macaque PBMCs. An analysis of chimeric viruses constructed between the variant SIVMne170 and the parent virus SIVMneCL8 demonstrated that there are determinants encoded within both the 5' and 3' halves of SIVMne170 that independently contribute to its rapid-high/SI phenotype. As we previously observed with other SIVMne variants, the Env SU of SIVMne170 was important for syncytium induction but was not a key determinant of cytopathicity. By contrast, the intracellular domain of the envelope transmembrane protein (Env TM) contributed to both the SI and cytopathic properties of SIVMne170. We also found that the minimal determinant within the 5' half of SIVMne170 that conferred its rapid replication kinetics and cytopathicity mapped to the capsid- and nucleocapsid-encoding regions of gag. Together, these data demonstrate that mutations selected in Gag and Env TM intracytoplasmic tail influence the replication and cytopathicity of SIVMne variants that evolve in the host.     

Mutational analysis of the human immunodeficiency virus vpr open reading frame.

Mutations were introduced by recombinant DNA techniques into the vpr open reading frame of an infectious molecular clone of human immunodeficiency virus type 1. The effect of these changes on the replicative and cytopathologic properties of the virus recovered from transfected cells was studied in several human CD4+ lymphocyte cell lines. In all cases, mutant viruses were infectious and cytopathic. However, when a low-input dose was used, mutants grew significantly more slowly than the wild-type virus. The growth kinetics of vpr mutants were distinct from those of vif and vpu mutants.     

Immunoblot analysis of trypomastigote excreted-secreted antigens as a tool for the characterization of Trypanosoma cruzi strains and isolates

An analysis of antibody recognition of Trypanosoma cruzi exoantigens by immunoblotting revealed a unique banding pattern that seems to be characteristic of each strain or isolate. Trypomastigote excreted-secreted antigens (TESA) present in supernatants of LLC-MK2 cells infected with 5 strains and 10 isolates of T. cruzi produced 13 different immunoblotting patterns. The same bands were observed when probed with acute-phase Chagas' disease serum or with serum from a rabbit immunized with the repetitive domain of T. cruzi transialidase recombinant protein (anti-shed acute-phase antigens). Three similar patterns were observed with TESA from 3 human isolates that probably belong to the same T. cruzi strain. When clone CL Brener, clone CL-14, and CL parental strain were analyzed, the same bands were observed, although they presented different biological behavior. These results suggest that immunoblotting analysis of TESA may be a useful tool for characterization of T. cruzi strains and isolates.     

Cellular origin of histamine-releasing factor produced by peripheral blood mononuclear cells

Histamine releasing factors (HRF) are a group of cytokines that release histamine and other mediators from mast cells and basophils. It has been speculated that HRF might play a major role in the pathogenesis of allergic diseases. Most investigators have studied PBMC as a source of HRF. This study was undertaken to investigate the cellular origin of HRF. Peripheral blood was processed to isolate and purify monocytes, T cells, CD4- T cells, CD8- T cells and B cells by using plastic adherence, 2-aminoethylisothiomonium-treated SRBC rosetting and negative selection with the use of mAb OKM1, OKT11, OKT8, OKT4, and OKB7 plus C. Highly purified subpopulations of PBMC were cultured alone or in the presence of Con A for 24 h. Supernatants were harvested, dialyzed, and assayed for HRF activity in the basophil histamine release test. We found that all subpopulations of PBMC including T cells, CD4- T cells, CD8- T cells, B cells, and monocytes produce variable quantities of HRF. The spontaneous production is very high in B cells but only barely measurable in T cells and monocytes. The synthesis of HRF by B cells was confirmed by abolishing the release of the activity after treatment of B cells with OKB7 mAb and C. Stimulation of cell populations by Con A significantly enhances HRF production by PBMC and T cells but not by B cells and monocytes. In mixing experiments, unstimulated monocytes + B cells showed synergism, but other combinations demonstrated an additive effect. This is the first demonstration of HRF production by human peripheral blood B cells. The results of this study also suggest that histamine releasing cytokines are of multiple cellular origin. This perhaps contributes to their molecular heterogeneity.     

Primary Human Leukocyte Subsets Differentially Express Vaccinia Virus Receptors Enriched in Lipid Rafts

Poxviruses, including vaccinia virus (VV) and canarypox virus (ALVAC), do not indiscriminately infect all cell types of the primary human leukocytes (PHLs) that they encounter but instead demonstrate an extremely strong bias toward infection of monocytes and monocyte lineage cells. We studied the specific molecular events that determine the VV tropism for major PHL subsets including monocytes, B cells, neutrophils, NK cells, and T cells. We found that VV exhibited an extremely strong bias of cell surface protein-dependent binding to monocytes, B cells, and activated T cells to a similar degree and to neutrophils to a much lesser extent. Resting T cells and resting NK cells exhibited only trace amounts of VV binding. Activated T cells, however, became permissive to VV binding, infection, and replication, while activated NK cells still resisted VV binding. VV binding strongly colocalized with lipid rafts on the surfaces of all VV binding-susceptible PHL subsets, even when lipid rafts were relocated to cell uropods upon cell polarization. Immunosera raised against detergent-resistant membranes (DRMs) from monocytes or activated T cells, but not resting T cells, effectively cross-blocked VV binding to and infection of PHL subsets. CD29 and CD98, two lipid raft-associated membrane proteins that had been found to be important for VV entry into HeLa cells, had no effect on VV binding to and infection of primary activated T cells. Our data indicate that PHL subsets express VV protein receptors enriched in lipid rafts and that receptors are cross-presented on all susceptible PHLs.     

Distinctive features in the production of IL-6 by human T cells

The production of IL-6 was studied in purified populations of human peripheral blood T cells and monocytes. IL-6 could be elicited from both T cells and thymocytes by a variety of polyclonal stimuli. The expression and production of IL-6 in T cells varied in several ways from that of monocytes: (1) under maximal stimulation, T cells secreted 10-fold less IL-6 than did monocytes; (2) by Western blot, the molecular isoforms produced by T cells were distinct from those produced by monocytes, with at least some isoforms being unique to each cell type; (3) the kinetics of IL-6 expression and production was much slower in T cells than in monocytes; (4) the frequency of labeled cells detected by in situ hybridization was 4% for T cells and 34% for monocytes. Our data indicate that there is considerable diversification among cell types in terms of regulation of IL-6 production and possibly in post-translational modifications. This raises the possibility that the IL-6 produced by different cell types may have varying effects depending on the form produced and the timing of synthesis.