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Human herpesvirus 6 (HHV-6) is a lymphotropic herpesvirus, and in vitro, HHV-6 can productively infect many of the same cell types as can human immunodeficiency virus (HIV). Coinfection by both viruses in vitro can lead to both activation of the HIV promoter and acceleration of cytopathic effects. We have previously demonstrated that a large, 22.25-kb cloned HHV-6 fragment, pZVB70, can trans activate HIV promoter expression in vitro. In this study, we show that the pZVB70 fragment can trans activate the HIV promoter in human T-cell lines as well as in the monkey kidney cell line CV-1. The pZVB70 insert was digested with various restriction enzymes, and individual fragments were transfected into cells to test for their ability to trans activate the HIV promoter. By this method, we have identified a 1.8-kb subfragment, B701, that is involved in trans activation. Sequence analyses show that B701 potentially encodes a 143-amino-acid protein. This protein shares no homology with other herpesvirus proteins, such as ICP0 and ICP4, that have been shown to trans activate the HIV promoter. However, it shows weak sequence homology with the gene products encoded by the cytomegalovirus early US22 gene family, suggesting that the putative B701 protein may be an HHV-6 early regulatory protein. The 143-amino-acid coding sequence of B701 was cloned by polymerase chain reaction, and transfection of this construct into cells activated HIV promoter expression. The target site on the HIV promoter for the putative B701 protein is mapped to the NF-kappa B binding site. Our results suggest that the putative B701 protein may function by directly binding to the NF-kappa B site or may involve cellular factors, such as NF-kappa B, either directly or indirectly.

The value of beta 2-microglobulin and neopterin concentrations in serum for early diagnosis of infants born to human immunodeficiency virus type 1 (HIV-1)-infected mothers was assessed. Concentrations of both markers were measured in serum samples from pediatric patients (Centers for Disease Control classifications P0, P1, and P2), as well as in age-matched normal subjects. Both beta 2-microglobulin and neopterin were significantly increased in HIV-1-infected symptomatic subjects (P2) compared to controls. Seventy-five percent of asymptomatic patients (P1) also had increased values. On the other hand, a significant overlap in concentrations of both markers in serum was found between controls and P0 patients. Thirty-eight percent of the P0 patients had values comparable to those of the P2 group. Persistently high concentrations of both markers in P0 patients may be indicative of HIV-1 infection.

The mechanism of trans-acting regulation of transcription from the long terminal repeat (LTR) of human immunodeficiency virus (HIV) has been investigated. The roles of the cis-acting elements within HIV LTR, as well as the trans-acting factors present in HIV-infected cells, have been evaluated by an in vitro transcription system. Our observations indicate that both the sequence downstream from the CAP site of HIV LTR (located at nucleotide positions +1 to +56; called the TAR element) and the GC boxes (-77 to -45) are required for full transcriptional stimulation and that both the virus-encoded tat protein and one or more cellular factors might be involved. These results demonstrate the presence of a combinatorial regulation of HIV transcription by multiple factors, which may confer the provirus with greater flexibility in regulated viral gene expression.

We have shown that 6D5 cells infected with the HIV-1 strain HTLV-III451 (6D5(451)) secreted viral envelope proteins gp160 and gp120 into the culture medium. Single cell cloning of 6D5(451) cells separated three distinct phenotypes. All clones secreted unprocessed env protein gp160 along with gp120. Only one phenotype produced infectious virus and contained normally processed gag proteins. The second phenotype was associated with nonproducer cells expressing only the env gene but no extracellular particles. The third phenotype synthesized Pr53gag but no reverse transcriptase, nor did it process the gag precursor. Only immature particles could be seen in the culture. Cells of the first and the third phenotypes produced two sizes of gp160, the normal and one with a small truncation at the C-terminus. Phenotype 2 only produced the smaller gp160. In all cases the gp160 that was secreted into the medium was the truncated molecule.

Histologic studies supplemented by in situ hybridization for human immunodeficiency virus type 1 (HIV-1), Epstein-Barr virus, cytomegalovirus, and human herpesvirus type 6 were performed on tissues obtained from the autopsy of six patients who died either by homicide or suicide shortly after learning of their seroconversion. Except for mild nonspecific lymphoid tissue reactions, no lesions were noted that would indicate HIV-1 infection. DNA from all viruses was detected in some lymphoid cells. The amount of DNA for Epstein-Barr virus, cytomegalovirus, and human herpesvirus type 6 corresponded to that observed for clinically occult latent infection. Lymphoid cells carrying HIV-1 DNA were even less frequent. Cells positive for HIV-1 were noted in the lamina propria of the large intestine in three male homosexuals and in one female prostitute. The cells were arranged similar to antigen-presenting cells. The present findings are consistent with current theories regarding the pathogenesis of HIV-1-associated disease.

We have previously shown that the presence of the EBV genome and active EBV infection in some Burkitt's lymphoma cell lines confer the susceptibility of HIV-1 infection in spite of the absence of surface CD4 receptors in these cells. In the present study we used an EBV genome negative Burkitt's lymphoma B-cell line, DG75, transfected with three different subgenomic fragments of EBV expressing the nuclear antigens (EBNA1 and EBNA2) and the latent membrane protein (LMP), respectively. Immunofluorescence analysis demonstrated CD4 expression in more than 90% of the EBNA1, EBNA2 and LMP transfected cell lines, whereas the antigen could only be detected in less than 4% of the parental DG75 cell line. Unlike the wild type DG75 line, the three transfected cell lines were shown to be susceptible to HIV-1 by both IFA and production of virions. Northern blotting of poly(A) selected mRNA of the four cell lines and hybridization to a human CD4 cDNA (pT4B) demonstrated a 3 kb band in all three EBNA1, EBNA2 and LMP transfected cells as well as in the wild type DG75 cells. Approximate quantification indicates equivalent level of T4 mRNA expression in the transfected cell lines as compared to T-cell lines (Hut-78, H9-9).

The long terminal repeats (LTRs) of the human immunodeficiency virus (HIV) the Rous sarcoma virus (RSV) and the copia Drosophila retrotransposon were compared in their capacity to direct expression of the bacterial cat (chloramphenicol acetyltransferase) gene in human, murine, and Drosophila cell lines. The results indicate that HIV and RSV LTR expression is post transcriptionally repressed in the Drosophila cells while copia LTR expression is post-transcriptionally repressed in the human and murine cells.

Human Herpes virus-6 (HHV-6) can co-infect with HIV-1 human CD4+ T-cells, leading to accelerated cell death, and factors in HHV-6-infected cells stimulate HIV-1 LTR directed gene expression. In this study, we have examined the mechanism of HIV-1 activation by HHV-6 and localized the cis-acting sequences of HIV-1 LTR responsive to trans-activation. Increased HIV-1 LTR directed gene expression is obtained in HIV-1 infected cells co-infected with HHV-6, or in HHV-6 infected cells co-transfected with the HIV-1 tat gene. Parallel increases of HIV-1-specific transcripts are seen by in situ hybridization in HHV-6/HIV-1 doubly infected cells as compared to single HIV-1 infection. Similarly, infection by HHV-6 increases the steady-state level of HIV-1 LTR mRNA that parallels CAT enzymatic activity, suggesting a transcriptional and/or post-transcriptional activation. Sequences necessary for HIV-1 LTR activation by HHV-6 are distinct from those required for that tat response and map to a region of the HIV-1 LTR from -103 to -48. The HIV-1 enhancer sequence (-105 to -80) is sufficient to confer HHV-6 inducibility to a heterologous promoter, and nuclear protein(s) activated or induced by HHV-6 infection specifically bind to the NF kappa B motifs of the HIV-1 enhancer region.(ABSTRACT TRUNCATED AT 250 WORDS)

It is now well established that cytokines are involved in the regulation of gene expression from HIV-1 LTR. The present study provides evidence that TNF-alpha stimulates HIV-1 gene expression and that the enhancer sequence within the HIV-1 LTR is involved in the stimulation. These results support the idea that immunologic stimulation and infection may trigger the development of clinical AIDS in individuals latently infected with HIV-1.