STAT1: A Novel Target of miR-150 and miR-223 Is Involved in the Proliferation of HTLV-I–Transformed and ATL Cells

AbstractWe have previously reported on the deregulation of cellular microRNAs involved in hematopoiesis and inflammation in human T-cell lymphotropic virus type 1 (HTLV-I)–transformed cells. In this study, we demonstrate that miR-150 and miR-223 specifically target the signal transducer and activator of transcription 1 (STAT1) 3′ untranslated region, reducing STAT1 expression and dampening STAT1-dependent signaling in human T cells. The effects of miR-150 and miR-223 on endogenous STAT1 were confirmed using inducible cell lines. Our studies also showed that miR-150 expression is upregulated by interleukin-2 signaling in adult T cell leukemia/lymphoma (ATL) cells. HTLV-I–transformed and ATL-derived cells have reduced levels of miR150 and miR223 expression, which coincide with increased STAT1 expression and STAT1-dependent signaling. Knockdown of STAT1 by short hairpin RNA demonstrated that the constitutive activation of STAT1 is required for the continuous proliferation of HTLV-I–transformed cells. Our studies further demonstrate that increased expression of STAT1 in ATL cells is associated with higher levels of major histocompatibility complex class I expression. Previous studies have demonstrated that the pressure exerted by natural killer (NK) cells in vivo can edit leukemic tumor cells by forcing an increased expression of major histocompatibility complex class I to escape immune clearance. STAT1-expressing tumor cells produce more aggressive tumors because they cannot be eliminated by NK cells. Our results suggest that therapeutic approaches using combined targeting of STAT1 and MHC class I may be an effective approach to activate NK cell–mediated clearance of ATL tumor cells

Genome wide analysis of human genes transcriptionally and post-transcriptionally regulated by the HTLV-I protein p30

<p>Abstract</p> <p>Background</p> <p>Human T-cell leukemia virus type 1 (HTLV-I) is a human retrovirus that is etiologically linked to adult T-cell leukemia (ATL), an aggressive and fatal lymphoproliferative disease. The viral transactivator, Tax, is thought to play an important role during the initial stages of CD4⁺T-cell immortalization by HTLV-1. Tax has been shown to activate transcription through CREB/ATF and NF-KB, and to alter numerous signaling pathways. These pleiotropic effects of Tax modify the expression of a wide array of cellular genes. Another viral protein encoded by HTLV-I, p30, has been shown to affect virus replication at the transcriptional and posttranscriptional levels. Little is currently known regarding the effect of p30 on the expression and nuclear export of cellular host mRNA transcripts. Identification of these RNA may reveal new targets and increase our understanding of HTLV-I pathogenesis. In this study, using primary peripheral blood mononuclear cells, we report a genome wide analysis of human genes transcriptionally and post-transcriptionally regulated by the HTLV-I protein p30.</p> <p>Results</p> <p>Using microarray analysis, we analyzed total and cytoplasmic cellular mRNA transcript levels isolated from PBMCs to assess the effect of p30 on cellular RNA transcript expression and their nuclear export. We report p30-dependent transcription resulting in the 2.5 fold up-regulation of 15 genes and the down-regulation of 65 human genes. We further tested nuclear export of cellular mRNA and found that p30 expression also resulted in a 2.5 fold post-transcriptional down-regulation of 90 genes and the up-regulation of 33 genes.</p> <p>Conclusion</p> <p>Overall, our study describes that expression of the HTLV-I protein p30 both positively and negatively alters the expression of cellular transcripts. Our study identifies for the first time the cellular genes for which nuclear export is affected by p30. These results suggest that p30 may possess a more global function with respect to mRNA transcription and the nuclear shuttling of cellular mRNA transcripts. In addition, these alterations in gene expression may play a role in cell transformation and the onset of leukemia.</p

HTLV-I p30 inhibits multiple S phase entry checkpoints, decreases cyclin E-CDK2 interactions and delays cell cycle progression

<p>Abstract</p> <p>Background</p> <p>Human T-cell leukemia virus type I (HTLV-I) has efficiently adapted to its host and establishes a persistent infection characterized by low levels of viral gene expression and slow proliferation of HTLV-I infected cells over decades. We have previously found that HTLV-I p30 is a negative regulator of virus expression.</p> <p>Results</p> <p>In this study we show that p30 targets multiple cell cycle checkpoints resulting in a delayed entry into S phase. We found that p30 binds to cyclin E and CDK2 and prevents the formation of active cyclin E-CDK2 complexes. In turn, this decreases the phosphorylation levels of Rb and prevents the release of E2F and its transcriptional activation of genes required for G1/S transition. Our studies also show that HTLV-II p28 does not bind cyclin E and does not affect cell cycle progression.</p> <p>Conclusions</p> <p>In contrast to HTLV-I, the HTLV-II-related retrovirus is not oncogenic in humans. Here we report that the HTLV-I p30 delays cell cycle progression while its homologue, HTLV-II p28, does not, providing evidence for important differences between these two related retrovirus proteins.</p

Germinal epimutation of Fragile Histidine Triad (FHIT) gene is associated with progression to acute and chronic adult T-cell leukemia diseases

Background: Human T cell Leukemia virus type 1 (HTLV-I) is etiologically linked to adult T cell leukemia/lymphoma (ATL) and an inflammatory neurodegenerative disease called HTLV-I-associated myelopathy or tropical spastic paraparesis (HAM/TSP). The exact genetic or epigenetic events and/or environmental factors that influence the development of ATL, or HAM/TSP diseases are largely unknown. The tumor suppressor gene, Fragile Histidine Triad Diadenosine Triphosphatase (FHIT), is frequently lost in cancer through epigenetic modifications and/or deletion. FHIT is a tumor suppressor acting as genome caretaker by regulating cellular DNA repair. Indeed, FHIT loss leads to replicative stress and accumulation of double DNA strand breaks. Therefore, loss of FHIT expression plays a key role in cellular transformation. Methods: Here, we studied over 400 samples from HTLV-I-infected individuals with ATL, TSP/HAM, or asymptomatic carriers (AC) for FHIT loss and expression. We examined the epigenetic status of FHIT through methylation specific PCR and bisulfite sequencing; and correlated these results to FHIT expression in patient samples. Results: We found that epigenetic alteration of FHIT is specifically found in chronic and acute ATL but is absent in asymptomatic HTLV-I carriers and TSP/HAM patients' samples. Furthermore, the extent of FHIT methylation in ATL patients was quantitatively comparable in virus-infected and virus non-infected cells. We also found that longitudinal HTLV-I carriers that progressed to smoldering ATL and descendants of ATL patients harbor FHIT methylation. Conclusions: These results suggest that germinal epigenetic mutation of FHIT represents a preexisting mark predisposing to the development of ATL diseases. These findings have important clinical implications as patients with acute ATL are rarely cured. Our study suggests an alternative strategy to the current "wait and see approach" in that early screening of HTLV-I-infected individuals for germinal epimutation of FHIT and early treatment may offer significant clinical benefits

Small PARP inhibitor PJ-34 induces cell cycle arrest and apoptosis of adult T-cell leukemia cells

A grant from the One-University Open Access Fund at the University of Kansas was used to defray the author’s publication fees in this Open Access journal. The Open Access Fund, administered by librarians from the KU, KU Law, and KUMC libraries, is made possible by contributions from the offices of KU Provost, KU Vice Chancellor for Research & Graduate Studies, and KUMC Vice Chancellor for Research. For more information about the Open Access Fund, please see http://library.kumc.edu/authors-fund.xml.Background HTLV-I is associated with the development of an aggressive form of lymphocytic leukemia known as adult T-cell leukemia/lymphoma (ATLL). A major obstacle for effective treatment of ATLL resides in the genetic diversity of tumor cells and their ability to acquire resistance to chemotherapy regimens. As a result, most patients relapse and current therapeutic approaches still have limited long-term survival benefits. Hence, the development of novel approaches is greatly needed. Methods In this study, we found that a small molecule inhibitor of poly (ADP-ribose) polymerase (PARP), PJ-34, is very effective in activating S/G2M cell cycle checkpoints, resulting in permanent cell cycle arrest and reactivation of p53 transcription functions and caspase-3-dependent apoptosis of HTLV-I-transformed and patient-derived ATLL tumor cells. We also found that HTLV-I-transformed MT-2 cells are resistant to PJ-34 therapy associated with reduced cleaved caspase-3 activation and increased expression of RelA/p65. Conclusion Since PJ-34 has been tested in clinical trials for the treatment of solid tumors, our results suggest that some ATLL patients may be good candidates to benefit from PJ-34 therapy