Guanylate-binding proteins (GBPs) are the most abundant cellular proteins expressed in response to interferon-gamma (IFN-gamma), with seven highly homologous members in humans, termed HuGBP-1 to HuGBP-7. To date, differential features that may indicate differential functions of these proteins have not been described. Here, we investigated the expression and subcellular localization of the different HuGBPs in endothelial cells (EC). IFN-gamma, tumor necrosis factor-alpha (TNF-alpha), and interleukin-1beta (IL-1beta) induced the expression of HuGBP-1, HuGBP-2, and HuGBP-3 at similar high levels. In contrast, expression of HuGBP-4 and HuGBP-5 was robustly induced only by IFN-gamma and not by TNF-alpha and IL-1beta. Expression of HuGBP-6 and HuGBP-7 was not detected in EC under the various conditions examined. Investigating subcellular localization of the EC-expressed HuGBPs, HuGBP-1, HuGBP-3, and HuGBP-5 were exclusively detected in the cytoplasm, whereas HuGBP-2 and HuGBP-4 displayed a nucleocytoplasmic distribution. Treatment of the cells with IFN-gamma and aluminum fluoride caused rapid enrichment of HuGBP-1 and HuGBP-2 in the Golgi apparatus, as demonstrated by time-lapse microscopy and fluorescence analyses of GFP-tagged HuGBPs. HuGBP-3 and HuGBP-4 were never detected in the Golgi apparatus, whereas HuGBP-5 was constitutively enriched in this cytosolic compartment, irrespective of stimulation. These results assign a characteristic pattern of expression and subcellular localization to each of the HuGBPs, indicating for the first time that these proteins may have different cellular functions.

Human herpesvirus 8 (HHV-8

Herpesvirus saimiri C488 transforms human T lymphocytes to stable growth in culture. The growth-transformed human T cells harbor the viral genome in a nonintegrated episomal form without production of virus particles. In these cells, virus gene expression was previously found to be confined to the transforming genes stpC and tip. In order to analyze virus gene expression in more detail, we applied a subtractive hybridization technique and compared stimulated virus-transformed cells with uninfected parental T cells of the same donor. A number of known T-cell activation genes were isolated. Viral stpC/tip cDNAs were enriched after subtraction. In addition, the viral immediate-early, superantigen-homologous gene ie14/vsag was represented by numerous cDNA clones that comprised the entire spliced transcript. Whereas a weak basal expression of ie14/vsag was detected by reverse transcription-PCR only, the phorbol ester-induced transcripts were readily shown by Northern blotting. ie14/vsag, which before had been classified as a major immediate-early gene of herpesvirus saimiri, is localized within a highly conserved region with extensive homologies to the cellular genome. Mutant viruses without the ie14/vsag gene are replication competent and fully capable of transforming human and marmoset T cells. Since ie14/vsag is transiently expressed after stimulation, it may increase T-cell proliferation in an activation-dependent and superantigen-like but apparently V␤-independent way.

Herpesvirus saimiri (HVS), the rhadinovirus prototype, is apathogenic in the persistently infected natural host, the squirrel monkey, but causes acute T cell leukemia in other New World primate species. In contrast to subgroups A and B, only strains of HVS subgroup C such as C488 are capable of transforming primary human T cells to stable antigen-independent growth in culture. Here, we report the complete 155-kb genome sequence of the transformation-competent HVS strain C488. The A+T-rich unique L-DNA of 113,027 bp encodes at least 77 open reading frames and 5 URNAs. In addition to the viral oncogenes stp and tip, only a few genes including the transactivator orf50 and the glycoprotein orf51 are highly divergent. In a series of new primary HVS isolates, the subgroup-specific divergence of the orf50/orf51 alleles was studied. In these new isolates, the orf50/orf51 alleles of the respective subgroup segregate with the stp and/or tip oncogene alleles, which are essential for transformation.

Human herpesvirus 8 (HHV-8