A retrovirus (ATLV) was unequivocally demonstrated in human adult T-cell leukemia (ATL) cell lines by density (1.152-1.155 g/cm3) in a sucrose gradient, reverse transcriptase activity insensitive to actinomycin D, RNA labeled with [3H]uridine, and specific proteins with molecular weights of 11,000, 14,000, 17,000, 24,000, and 45,000. Furthermore, cDNA prepared by endogenous reaction with detergent-treated virions hybridized to 35S RNA containing poly(A), which was inducible by IdUrd treatment of a T-cell line derived from leukemic cells of the ATL, and the integrated form of ATLV proviral DNA was detected in T-cell lines derived from ATL. The ATLV proviral DNA was also detected in fresh peripheral lymphocytes from all five patients with ATL tested so far but not in those from healthy adults. On the other hand, ATLV protein of Mr 42,000 was found to be at least one of the ATL-associated antigen(s) that were previously detected in ATL-leukemic cells by all sera from patients with ATL. These findings on the close association of ATLV protein and proviral DNA with ATL are direct evidence for the possible involvement of the retrovirus ATLV in leukemogenesis of human AUL.Adult T-cell leukemia (ATL) is a clinical entity of T-cell malignancy proposed by Takatsuki and colleagues (1, 2) on the basis of its characteristic clinical and hematological features. The disease has also been called "endemic adult leukemia/lymphoma (ATLL)" because of its leukemic lymphomatous nature and the peculiar geographic distribution ofthe birthplaces ofpatientsclustering in the southwestern part ofJapan (3). Recently, Hinuma et aL (4) found antigen(s) in a T-cell line, MT-1, derived from peripheral leukemia cells of a patient with ATL. The antigen(s) reacted with sera from all ATL patients tested and also with sera from about 25% of the healthy adults in the endemic area but with very few sera of subjects from nonendemic areas. Virus-like particles with type C appearance were also found by electron microscopy (4). These observations suggested the presence and possible relationship oftype C virus with ATL. Similar antigen(s) and type C particles were also detected in another T-cell line, MT-2, established from normal cord lymphocytes cocultivated with leukemia cells from an ATL patient (5) and reverse transcriptase activity was detected in the culture fluid of MT-2 cells (unpublished data).In this work, we have unequivocally demonstrated and characterized the ATL retrovirus (ATLV) by biochemical techniques in two T-cell lines, MT-1 and MT-2. Furthermore, we obtained data indicating a close association of ATLV proviral DNA with human ATL. These pieces of direct evidence suggest the involvement of ATLV in human leukemogenesis.MATERIALS AND METHODS Cells. Two cell lines, MT-1 and MT-2, were used. The MT-1 cell line (6) was derived from leukemia cells of peripheral blood from a patient with ATL, and the MT-2 cell line (5) was established from cord lymphocytes that had been cocultivated with leukemia cells from a patient with ATL. Both ar...

Human retrovirus adult T-cell leukemia virus (ATLV) has been shown to be closely associated with human adult T-cell leukemia (ATL) [Yoshida, M., Miyoshi, I. & Hinuma, Y. (1982) Proc. Natl. Acad. Sci. USA 79, 2031-2035]. The provirus of ATLV integrated in DNA of leukemia T cells from a patient with ATL was molecularly cloned and the complete nucleotide sequence of 9,032 bases of the proviral genome was determined. The provirus DNA contains two long terminal repeats (LTRs) consisting of 755 bases, one at each end, which are flanked by a 6-base direct repeat of the cellular DNA sequence. The nucleotides in the LTR could be arranged into a unique secondary structure, which could explain transcriptional termination within the 3' LTR but not in the 5' LTR. The nucleotide sequence of the provirus contains three large open reading frames, which are capable of coding for proteins of 48,000, 99,000, and 54,000 daltons. The three open frames are in this order from the 5' end of the viral genome and the predicted 48,000-dalton polypeptide is a precursor of gag proteins, because it has an identical amino acid sequence to that of the NH2 terminus of human T-cell leukemia virus (HTLV) p24. The open frames coding for 99,000- and 54,000-dalton polypeptides are thought to be the pol and env genes, respectively. On the 3' side of these three open frames, the ATLV sequence has four smaller open frames in various phases; these frames may code for 10,000-, 11,000-, 12,000-, and 27,000-dalton polypeptides. Although one or some of these open frames could be the transforming gene of this virus, in preliminary analysis, DNA of this region has no homology with the normal human genome.

We developed a novel promoter system, designated SRa, which is composed of the simian virus 40 (SV40) early promoter and the R segment and part of the U5 sequence (R-U5') of the long terminal repeat of human T-cell leukemia virus type 1. The R-U5' sequence stimulated chloramphenicol acetyltransferase (CAT) gene expression only when placed immediately downstream of the SV40 early promoter in the sense orientation. The SRoa expression system was 1 or 2 orders of magnitude more active than the SV40 early promoter in a wide variety of cell types, including fibroblasts and lymphoid cells, and was capable of promoting a high level of expression of various lymphokine cDNAs. These features of the SRoa promoter were incorporated into the pcD-cDNA expression cloning vector originally developed by Okayama and Berg.

The genome of human T-cell leukemia virus (HTLV) was surveyed in fresh tumor cells of 163 patients with lymphoma and leukemia from the southwest part of Japan where adult T-cell leukemia (ATL) is endemic. Leukemic cells of all 88 cases of ATL tested so far were found to contain the provirus genome and also found to be monoclonal with respect to the integration site of provirus genome. In most cases of ATL, leukemic cells contained one or two copies of the complete HTLV provirus genome, and it was shown that the single species of HTLV with a fully determined sequence is typical in ATL. Some cases of T-cell malignancies, diagnosed as chronic lymphocytic leukemia or non-Hodgkin lymphoma, also had the provirus genome in their tumor cells, whereas some cases with the same diagnosis did not. No cases of other types of lymphoma or leukemia contained the provirus genome in their tumor cells. Monoclonal integration of the HTLV provirus genome in all primary tumor cells of ATL not only indicates that HTLV directly interacts with target cells, which become leukemic, and that integration of the provirus genome is a prerequisite for development of ATL and possibly other related diseases but also indicates that the virus is not associated with other types of lymphoma or leukemia.Human retroviruses HTLV (human T-cell leukemia virus) (1-3) and ATLV (adult T-cell leukemia virus) (4, 5) were independently isolated from cases of cutaneous T-cell lymphoma (CTCL) and adult T-cell leukemia (ATL) (6), respectively. Subsequently, HTLV and ATLV were shown to be similar in immunological crossreactivities (7,8) and nucleic acid hybridization (9). Recently, we determined the total nucleotide sequence of the ATLV provirus genome (10) and showed that ATLV and HTLV type I are identical with respect to the locations of gene-specific sequences and the sites of some restriction enzymes (11). We use the term ATK strain of HTLV (HTLVATK) for the virus previously cloned in ATK-1 DNA and reported as ATLV (5, 10). The retrovirus HTLV is exogenous for humans (2, 5) and distinct from known animal retroviruses with respect to the structure of its provirus genome (10, 12). Furthermore, HTLV was shown to be closely associated with a unique T-cell malignancy, ATL, by extensive surveys of antibodies against the viral proteins (4, 7, 8, 13-15). The association of the virus with ATL was also shown by detecting the provirus genome in leukemic cells of ATL patients (5,16). Some patients with other types of lymphoma or leukemia, such as CTCL, chronic lymphocytic leukemia (T-CLL), and non-Hodgkin lymphoma of T-cell origin, were also reported to have antibodies against the viral proteins (4, 17). The presence of antibodies to viral proteins is evidence for infection with HTLV but does not provide any information on the mode of involvement of the virus in leukemogenesis.To understand whether HTLV is directly involved in leukemogenesis of ATL and whether the virus is associated with any other types of lymphomas or leukemias, we surveyed the provirus genome ...

The human T cell leukemia virus-1 (HTLV-1) is a retrovirus that causes adult T cell leukemia (ATL) and neurological disorder, the tropical spastic paraparesis (HAM/TSP). The pathogenesis apparently results from the pleiotropic function of Tax protein, which is a key regulator of viral replication. Tax exerts (a) trans-activation and -repression of transcription of different sets of cellular genes through binding to groups of transcription factors and coactivators, (b) dysregulation of cell cycle through binding to inhibitors of CDK4/6, and (c) inhibition of some tumor suppressor proteins. These effects on a wide variety of cellular targets seem to cooperate in promoting cell proliferation. This is an effective viral strategy to amplify its proviral genome through replication of infected cells; ultimately it results in cell transformation and leukemogenesis.