Immunological characterization of the low molecular weight gag gene proteins p19 and p15 of human T-cell leukemia-lymphoma virus (HTLV) and demonstration of human natural antibodies to them

Small molecular weight gag gene proteins p19 and p15 of human T-cell leukemia virus (HTLV) were purified to homogeneity from density banded virus, using differential affinities to phosphocellulose at pH 7.9 and 6.5. Using specific radioimmunoassays, p19 and p15 were shown to band with HTLV on sucrose gradients. These proteins were present only in HTLV-I and in cell lines expressing HTLV-I. They were not expressed in a B-cell line from patient CR, although his T cells were HTLV producers. No significant immunological cross-reactivity was detected between these proteins and HTLV-II_MO P19 and p15 did not share any common antigenic determinants with each other or with HTLV p24 in competition radioimmunoassays. Sera of HTLV-positive leukemia-lymphoma patients and some of their relatives also had natural antibodies to p19 and p15.     

Human T-cell leukemia virus type I isolates from Gabon and Ghana: comparative analysis of proviral genomes

Two isolates of human T-cell leukemia virus type I (HTLV-I) were obtained from lymphocyte cultures of a healthy carrier in Gabon and another in Ghana. Their proviruses were analyzed by Southern blot hybridization and compared with prototypical HTLV-I isolated in Japan and the United States. The provirus genomes of both strains were highly homologous to the prototype HTLV-I along the whole viral genome. The restriction endonuclease sites of the Ghanian isolate were almost identical with those of the prototype HTLV-I, but 10 of 26 sites of the Gabonese isolate were different from those of the prototype. Furthermore, the restriction map of the Gabonese isolate resembled those of a simian T-cell leukemia virus (STLV-I) isolated from a chimpanzee from Sierra Leone and a variant of HTLV-I from Zaire (HTLV-Ib) more closely than those of any other known HTLV-I. These results indicated the existence of some unique strains of HTLV-I transmitted among African people, and the importance of clarifying the origin and transmission of HTLV group viruses.     

HTLV type I (U. S. isolate) and ATLV (Japanese isolate) are the same species of human retrovirus

Two independent isolates of human leukemia virus, human T-cell leukemia virus (HTLV) and adult T-cell leukemia virus (ATLV), are shown to be the same by blotting analysis using gene-specific probes and restriction enzymes. Therefore, Japanese ATL virus and Caribbean HTLV type I, which are exogenous for human, have a common origin.     

Sequence homology of the simian retrovirus genome with human T-cell leukemia virus type I

A retrovirus highly related to human T-cell leukemia virus type I (HTLV-I) was isolated from a T-cell line established from a seropositive pig-tailed monkey and the provirus genome was molecularly cloned using HTLV-I as a probe. The monkey virus (STLV) had the genomic structure of the LTR-gag-pol-env-pX-LTR. Analysis of the env-pX-LTR region revealed the 90% homology of the nucleotide sequence with that of HTLV-I in each region. This high homology of the sequence indicates that STLV is a member of the HTLV family, but apparently different from HTLV-I. This suggests that the possibility of recent interspecies transmission from monkeys to humans in the endemic area is very small. From its similarity to HTLV, STLV should be useful as an animal model in studies on natural HTLV infection and leukemogenesis of HTLV in humans.     

Identification of HTLV p19 specific natural human antibodies by competition with monoclonal antibody

A competitive binding assay using a monoclonal antibody to the human T-cell lymphoma/leukemia virus (HTLV) p19 was developed for use in detecting natural antibodies to the protein in human sera. The specificity of the assay for HTLV p19 was demonstrated using a variety of antisera. While sera known to contain antibodies to HTLV p19 competed in the assay, antisera prepared against purified HTLV p24, the major core protein of the virus, or against other disrupted type-C retroviruses did not. Sera of Japanese patients with adult T-cell leukemia and similar T-cell malignant lymphomas were examined by this technique for the presence of antibodies to HTLV p19. The results were compared with those obtained by a solid-phase radioimmunoassay (RIA) against disrupted HTLV. The majority of Japanese ATL patients possess natural antibodies to HTLV as shown by solid-phase RIA (88%) and also specifically to HTLV p19 (77%). Similarly, 50% of Japanese patients with similar T-cell malignant lymphomas possess HTLV antibodies by solid-phase RIA and nearly as many (42%) possess anti-p19 reactivity. Twelve and eight percent, respectively, of normal Japanese donors from the ATL endemic region possessed HTLV-specific antibody by the solid-phase RIA or competitive binding assay. Normal donors from nonendemic areas lacked antibodies to HTLV. These results extend our previous findings of natural antibodies to HTLV in Japanese patients with ATL. The finding of p19-specific antibodies in these Japanese sera, together with previous reports of natural antibodies to HTLV p24 in sera from this same geographic cluster, strengthens the association of HTLV with Japanese ATL.     

Immune recognition of genetically diverse simian T-cell lymphotropic virus type I isolates

Summary Nucleotide sequence analysis of selected regions of the gag, pol, env and pX genes of simian T-cell lymphotropic virus type I (STLV-I) strains indicated that African isolates were more closely related to human T-cell lymphotropic virus type I (HTLV-I) than Asian isolates. Despite these recent comparative studies on nucleotide sequence homology between HTLV-I and STLV-I isolates, only limited information is available regarding the influence of genetic differences on antigen-antibody recognition of distinct STLV-I strains. In this study, we demonstrated that sera from STLV-I-infected yellow baboons (Papio cynocephalus) reacted strongly with env gp62/68 from HTLV-I-infected cell lines MT-2 and C10/MJ. In contrast, sera from Japanese macaques (Macaca fuscata) naturally infected with Asian STLV-I had weak reactivity to env gp62/68 of these prototypic HTLV-I strains. Pst-1 restriction enzyme analysis of proviral DNA indicated that the baboon virus isolates were more closely related to HTLV-I than the Japanese isolates. These results indicate that nucleotide sequence diversity, correlates with variations in proviral restriction enzyme sites and antibody recognition of viral envelope proteins. These differences in immunoreactivity may have important implications for serologic diagnosis, as well as epidemiological and vaccine studies of STLV-I infection.     

Human T-cell leukemia/lymphoma viruses. Life cycle, pathogenicity, epidemiology, and diagnosis.

Human T-cell leukemia/lymphoma virus type I (HTLV-I) was discovered in 1980, and it subsequently was found to be the cause of adult T-cell leukemia/lymphoma. A progressive neurologic disease known as tropical spastic paraparesis, or HTLV-I-associated myelopathy, has also been linked to infection with HTLV-I. A related virus, HTLV type II (HTLV-II), has been isolated from patients with hairy-cell leukemia, but it has not been proved to be the cause of any disease. In late 1988, US blood banks began screening all blood donations for antibodies to HTLV-I/II. This program has resulted in the identification of many unexpectedly seropositive blood donors and provided much information about the prevalence of HTLV-I/II in the United States. In this article, I review the replication of these agents, as well as their pathogenesis, diagnosis, and mechanisms of spread.     

Monoclonal antibodies against murine leukemia viruses: identification of six antigenic determinants on the p 15(E) and gp70 envelope proteins.

Hybrid cells that produced monoclonal antibodies against the envelope proteins of murine leukemia virus (MuLV) were prepared by the polyethylene glycol-mediated fusion of a mouse myeloma cell line with lymphocytes from mice immunized with allogeneic MuLV-producing leukemia cells. Twenty-three independent cell lines were cloned and inoculated into syngeneic mice for the production of ascites fluids that contained high-titered (20–75 mg/ml) monoclonal antibodies. Six serologically distinct specificities were detected when these ascites fluids were tested on a broad panel of MuLV and non-murine retra iruses. Prototype cell lines producing monoclonal antibodies that were representative of each pattern of reaction were selected for further study. In immune precipitation assays each of the prototype antibodies reacted with viral envelope proteins; three of these identified antigenic determinants on p15(E), while three others identified antigenic determinants on gp70. The p15(E) antigenic determinants were shared by a diverse panel of MuLV. One of these p15(E) antigenic determinants was also found in feline leukemia virus. The gp70 antigenic determinants, on the other hand, had a more restricted distribution and were found in only selected isolates of MuLV.     

Primary isolation of human T-cell leukemia-lymphoma virus types I and II: use for confirming infection in seroindeterminate blood donors.

We describe the use of an immunofluorescence assay and coculture to confirm human T-cell leukemia-lymphoma virus (HTLV) infection. Peripheral blood mononuclear cells from 32 of 32 seropositive donors were positive in the immunofluorescence assay, and 63% of their cocultures produced p24 antigen. Specific antibodies distinguished HTLV type I (HTLV-I) from HTLV-II. HTLV-I or HTLV-II was isolated from donors with indeterminate serologic test results.     

Glycosylation pattern of herpes simplex virus type 2 glycoprotein G from precursor species to the mature form

Summary Studies of adult T-cell leukaemia virus/human T-cell leukaemia/lymphotropic viruses (ATLV/HTLV-I) in Japan indicate that the virus is involved only with the development of ATL. By contrast, reports from the U.S.A. about HTLV have from time to time claimed that related HTLV are concerned not only with ATL of black persons, but also with a wide range of diseases, such as mycosis fungoides/Sezary's syndrome, T-cell hairy cell leukaemia, acquired immune deficiency syndrome (AIDS) and also multiple sclerosis. Using morphological, biological, serological and molecular hybridisation studies, we were able to confirm that the viruses implicated in the development of ATL and AIDS are distinct and that ATLV/HTLV-I is involved only in ATL, and HIV/LAV/HTLV-III only in AIDS. In vitro, ATLV/HTLV-I transformed and immortalised T-cells, while HIV/LAV/HTLV-III killed our T-cells. Failure to detect any serological cross-reaction indicates that all the structural proteins are different. Likewise, Southern blot studies failed to reveal any cross-hybridisation. Sixty patients with multiple sclerosis failed to reveal any association with ATLV/HTLV-I or with HIV/LAV/HTLV-III. Our conclusion is that ATLV/HTLV-I is involved only in ATL of Japanese and of some black persons of African origin, and that HIV/LAV/HTLV-III is associated only in AIDS.     

Monoclonal antibodies and chemiluminescence immunoassay for detection of the surface protein of human T-cell lymphotropic virus.

Monoclonal antibodies (MAbs) raised against human T-cell lymphotropic virus type I (HTLV-I) recognized five distinct antigenic domains of viral env gene-encoded proteins. By using recombinant env proteins and synthetic peptides as mapping antigens, it was determined that the most immunogenic region represented a central portion of the retroviral surface protein (domain 2; amino acids 165 to 191). However, only a single MAb was able to react strongly with native viral proteins. This antibody (clone 6C2) was directed to an epitope within domain 4 (amino acids 210 to 306) of the retroviral env gene and reacted with envelope proteins in both HTLV-I and HTLV-II, as determined by immunoprecipitation, solid-phase binding, and immunoblotting. No reactivity against envelope components of other human retroviruses, including human immunodeficiency virus types 1 and 2, was present. Flow cytometry data demonstrated that MAb 6C2 reacted with cell lines chronically infected with HTLV-I or HTLV-II and also with surface antigens expressed on fresh adult T-cell leukemia cells, following up-regulation with interleukin-2. By a chemiluminescence immunoassay procedure, picogram amounts of viral surface protein could be detected in the unconcentrated supernatants of HTLV-infected cell lines and in diagnostic cultures. Levels of env and gag proteins released by cells into culture supernatants were not directly related to percent expression of cell surface viral-coat proteins. Further, the molar ratio of p19 to gp46 in conditioned media varied from strain to strain, possibly reflecting differences in viral assembly or packaging mechanisms. MAb 6C2 will be of value in characterizing the biochemical and immunological behavior of retroviral env gene proteins and in studying the interaction of HTLV-I and HTLV-II with their receptors.     

Nucleotide sequence of the protease-coding region in an infectious DNA of simian retrovirus (STLV) of the HTLV-I family

A provirus clone of simian T-cell leukemia virus isolated from a pigtailed monkey (PT-STLV), which is 90% homologous to HTLV-I, was shown to be biologically active in transfection assay. In transfected cells, gp61env, Pr55gag, and the mature gag proteins p24, p21, and p15 were detected, and type C particles were produced. The virus could be transmitted from the transfectants to recipient cells by cocultivation. In this biologically active provirus clone, a coding frame, possibly for protease, was identified between the gag and pol genes. The corrected sequence of the protease region of HTLV-I was also found to have a single open reading frame overlapping the gag and pol genes, although it has an amber codon in the middle of the frame. Thus, a single coding frame, which is different from those of gag and pol, is common to proteases of the HTLV family including HTLV-I.     

The HTLV-I orfI protein is recognized by serum antibodies from naturally infected humans and experimentally infected rabbits

The mechanism of T-cell transformation by human T-cell lymphotropic virus type I (HTLV-I), though not completely understood, appears to involve the interactions of several viral and cellular proteins. One of these viral proteins, p12(I), encoded by HTLV-I orfI, is a weak oncogene that binds the 16-kDa subunit of the vacuolar ATPase and interacts with the immature beta and gamma(c) chains of the IL-2 receptor. We have expressed the singly spliced orfI cDNA in the baculovirus system and used the recombinant protein as a tool to assess the presence of antibodies in naturally or experimentally infected hosts. In addition, rabbit antisera were raised against various p12(I) synthetic peptides and used to identify three antigenic regions within p12(I), one between the two putative transmembrane regions of p12(I) and two at the carboxy-terminus of the protein. More importantly, sera from a naturally infected human (1 of 32) and experimentally infected rabbits (9 of 20) recognized the rp12(I), demonstrating orfI expression and immunogenicity in vivo. Taken together these data provide the first evidence of orfI expression during HTLV-I infections.