In 1980, Human T-cell leukemia virus type I (HTLV-1) became the first retrovirus discovered in humans. It has been revealed that infection by this virus causes diseases such as adult T-cell leukemia (ATL), HTLV-I associated myelopathy (HAM), and tropical spastic paraparesis (TSP), and that the symptoms of such diseases develop after a long period of time in the body, with an average latency of about 40 to 50 years after the infection with the virus. However, little is known about the mechanisms of its latent infection and onset.
The gene of HTLV-I includes a tax/rex region in addition to three regions which are common to animal retroviruses, i.e., a gag region, a pol region, and an env region. The tax/rex region, which is located down-stream of env, is considered to have an important role in the expression of viral gene and the onset of ATL. The mRNA of tax/rex results after double splicings from a primary transcript of the HTLV-I gene, and includes two overlapping open reading frames. From one open reading frame is translated a 40 kilodalton protein called Tax, which acts on the LTR of the virus itself as well as promoters of various genes in cells to activate transcription. From the other open reading frame is translated a 27 kilodalton protein called Rex, which controls the processing of the viral RNA occurring within the nucleus after transcription, and positively acts on the transport to the cytoplasm of unspliced mRNA.
From a different start codon within the same open reading frame as that of the Rex protein, a 21kD protein called p21X is translated. The inventors revealed that p21X is translated from the p21X mRNA which lacks the second exon through single splicing from the HTLV-I gene (Orita et al. FEBS Lett., 295, 127-134 (1991)). However, the functions of the protein are unknown. The inventors further discovered that the p21X mRNA is expressed by a mutant provirus having a deletion in a broad region encompassing the gag, pol, and env regions in a HTLV-I infected cell line (Orita et al. Nucleic Acids, Res., 21, 3799-3807 (1993)). Moreover, the inventors discovered that the p21X mRNA is also expressed by the above-mentioned mutant provirus in peripheral blood lymphocytes of patients infected with HTLV-I. On the other hand, it is known that the expression of mRNA for the tax/rex region from a complete provirus is rarely observed in the peripheral blood of patients infected with HTLV-I, and is detectable in vivo only by the RT-PCR method, which is an ultra-sensitive detection method. However, once peripheral blood lymphocytes of patients infected with HTLV-I are transferred to a culture system in vitro, its expression is known to become high enough to be easily detected (Kiyokawa et al., Proc. Natl. Acad. Sci. USA., 82, 8359-8363 (1985)). Moreover, the expression of the p21X mRNA has also been found to be on the same level before and after the culture of the aforementioned peripheral blood lymphocytes of patients infected with HTLV-I (Orita et al., J. Gen. Virol., 73, 2283-2289 (1992)). Therefore, it was considered that, in vivo, the p21X mRNA is expressed without being repressed, whereas the expression of the mRNA for tax/rex is repressed.
It is considered that HTLV-I and human immunodeficiency virus (HIV), which is a kind of RNA virus, delays splicing reactions by making the splicing signal within a DNA sequence a non-optimal one, thereby providing time for the Rex protein of HTLV-I or the Rev protein of HIV to repress splicing reactions (Chang, D. D. and Sharp. P. A., Science, 249, 614-615 (1990)). It is considered that sufficient amounts of the Rex protein and the Rev protein need to be expressed and accumulated within the nucleus in order to be polymerized, before their functions can be exhibited to promote the repression of the splicing of the viral mRNA having RXE (Rex responsible element) or RRE (Rev responsible element) and the transport to the cytoplasm, thereby triggering the replication of the viruses (expression of the structural proteins) (Inoue et al., Proc. Natl. Acad. Sci. USA., 84, 3653-3657 (1987); Hidaka et al., EMBO J., 7, 519-523 (1988); Seiki et al. Proc. Natl. Acad. Sci. USA., 85, 7124-7128(1988); and Hanly et al. Genes Dev., 3, 1534-1544(1989)).
The above indicates that at least two regulatory factors, i.e., Tax and Rex proteins, are translated from the pX region characteristic of HTLV-I, and that these factors are necessary for the replication of HTLV-I. Tax is a transcription activation factor which acts on the LTR (long terminal repeat) and also activates various cellular genes. Tax also has an ability to transform certain types of cultured cells. Thus, the possibility of Tax being involved in the oncogenesis of a cell by HTLV-I is suggested.
The expression of the HTLV-I gene is known to be very low in periods of inapparent infection and to be low even after the onset of ATL. Therefore, in order to elucidate the mechanism of latent infection of HTLV-I, it is considered important to study the expression repression mechanism of the viral gene. It is known that the expression control of the HTLV-I gene is performed mainly on the LTR of HTLV-1. The LTR region is subdivided into three regions called U3, R, and U5. The U3 region includes a sequence on which Tax acts, as well as sequences acted upon by CREB, ETS, AP1, etc., which are cellular transcription activation factors. The R region is known to include a sequence to which YB-1 binds to activate transcription (Kashanchi et al. J. Virol., 68(1): 561-565 (1994)). Moreover, the R and U5 regions include a region which functions repressively with respect to the HTLV-I gene expression on the transcription level or post-transcriptionally (Xu et al., Mol. Cell. Biol., 14(8): 5371-5383 (1994); and Seiki et al., Virology, 176: 81-86 (1990)). Furthermore, the inventors recently discovered a novel transcriptional repressive sequence (U5 repressive element; U5RE), and reported the existence of three proteins of 110 kDa, 80 kDa, and 70 kDa which specifically bind to U5RE (Okumura et al. FEBS Let., 356: 94-100 (1994)).
USPTO Backgrounds
USPTO Backgrounds