1: Mol Cell Biol. 2005 Jul;25(14):5869-79. Transcriptional repression of the Neurofibromatosis-1 tumor suppressor by the t(8;21) fusion protein. Yang G, Khalaf W, van de Locht L, Jansen JH, Gao M, Thompson MA, van der Reijden BA, Gutmann DH, Delwel R, Clapp DW, Hiebert SW. Department of Biochemistry, Vanderbilt University School of Medicine, PRB 512, 23rd and Pierce, Nashville, Tennessee 37232, USA. Von Recklinghausen's disease is a relatively common familial genetic disorder characterized by inactivating mutations of the Neurofibromatosis-1 (NF1) gene that predisposes these patients to malignancies, including an increased risk for juvenile myelomonocytic leukemia. However, NF1 mutations are not common in acute myeloid leukemia (AML). Given that the RUNX1 transcription factor is the most common target for chromosomal translocations in acute leukemia, we asked if NF1 might be regulated by RUNX1. In reporter assays, RUNX1 activated the NF1 promoter and cooperated with C/EBPalpha and ETS2 to activate the NF1 promoter over 80-fold. Moreover, the t(8;21) fusion protein RUNX1-MTG8 (R/M), which represses RUNX1-regulated genes, actively repressed the NF1 promoter. R/M associated with the NF1 promoter in vivo and repressed endogenous NF1 gene expression. In addition, similar to loss of NF1, R/M expression enhanced the sensitivity of primary myeloid progenitor cells to granulocyte-macrophage colony-stimulating factor. Our results indicate that the NF1 tumor suppressor gene is a direct transcriptional target of RUNX1 and the t(8;21) fusion protein, suggesting that suppression of NF1 expression contributes to the molecular pathogenesis of AML. PMID: 15988004 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 2: Endocr Relat Cancer. 2005 Jun;12(2):319-34. RET/PTC-induced gene expression in thyroid PCCL3 cells reveals early activation of genes involved in regulation of the immune response. Puxeddu E, Knauf JA, Sartor MA, Mitsutake N, Smith EP, Medvedovic M, Tomlinson CR, Moretti S, Fagin JA. Division of Endocrinology and Metabolism, University of Cincinnati College of Medicine, Cincinnati, Ohio 45267, USA. RET/PTC rearrangements represent key genetic events involved in papillary thyroid carcinoma (PTC) initiation. The aim of the present study was to identify the early changes in gene expression induced by RET/PTC in thyroid cells. For this purpose, microarray analysis was conducted on PCCL3 cells conditionally expressing the RET/PTC3 oncogene. Gene expression profiling 48 h after activation of RET/PTC3 identified a statistically significant modification of expression of 270 genes. Quantitative PCR confirmation of 20 of these demonstrated 90% accuracy of the microarray. Functional clustering of genes with greater than or less than 1.75-fold expression change (86 genes) revealed RET/PTC3-induced regulation of genes with key functions in apoptosis (Ripk3, Tdga), cell-cell signaling (Cdh6, Fn1), cell cycle (Il24), immune and inflammation response (Cxcl10, Scya2, Il6, Gbp2, Oas1, Tap1, RT1Aw2, C2ta, Irf1, Lmp2, Psme2, Prkr), metabolism (Aldob, Ptges, Nd2, Gss, Gstt1), signal transduction (Socs3, Nf1, Jak2, Cpg21, Dusp6, Socs1, Stat1, Stat3, Cish) and transcription (Nr4a1, Junb, Hfh1, Runx1, Foxe1). Genes coding for proteins involved in the immune response and in intracellular signal transduction pathways activated by cytokines and chemokines were strongly represented, indicating a critical role of RET/PTC3 in the early modulation of the immune response. PMID: 15947106 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 3: J Immunol. 2005 Apr 1;174(7):4144-52. Regulation of the murine Ddelta2 promoter by upstream stimulatory factor 1, Runx1, and c-Myb. Carabana J, Ortigoza E, Krangel MS. Department of Immunology, Duke University Medical Center, Durham, NC 27710, USA. Accessibility control of V(D)J recombination at Ag receptor loci depends on the coordinate activities of transcriptional enhancers and germline promoters. Recombination of murine Tcrd gene segments is known to be regulated, at least in part, by the Tcrd enhancer (Edelta) situated in the Jdelta2-Cdelta intron. However, there has been little characterization of promoters and other cis-acting elements that are activated by or collaborate with Edelta and that might function to regulate Tcrd gene recombination events. We now describe a strong promoter that is tightly associated with the murine Ddelta2 gene segment. EMSAs reveal that upstream stimulatory factor 1, Runx1, c-Myb, lymphoid enhancer binding factor 1, NF1, and E47 all interact with this promoter in vitro. Of these, upstream stimulatory factor 1, Runx1, and c-Myb appear necessary for full promoter activity in transiently transfected cells. Moreover, the same three factors were found to interact with the promoter in vivo by chromatin immunoprecipitation. We suggest that these factors play important roles as Edelta-dependent regulators of Ddelta2 accessibility in vivo. Consistent with the established roles of c-Myb and Runx factors in Edelta function, we detected low level, enhancer-independent activity of the Ddelta2 promoter in transient transfection experiments. We speculate that the Ddelta2 promoter may play a role as a weak, enhancer-independent regulator in vivo, and might contribute to residual Tcrd rearrangement in Edelta(-/-) mice. PMID: 15778374 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 4: J Virol. 1997 Oct;71(10):7273-80. Increased lymphomagenicity and restored disease specificity of AML1 site (core) mutant SL3-3 murine leukemia virus by a second-site enhancer variant evolved in vivo. Ethelberg S, Lovmand J, Schmidt J, Luz A, Pedersen FS. Department of Molecular and Structural Biology, University of Aarhus, Aarhus C, Denmark. SL3-3 is a highly T-lymphomagenic murine retrovirus. The major genetic determinant of disease is the transcriptional enhancer, which consists of a repeated region with densely packed binding sites for several transcription factors, including AML1 (also known as core binding factor and polyoma enhancer-binding protein 2) and nuclear factor 1 (NF1). Previously, we examined the enhancer structure of proviruses from murine tumors induced by SL3-3 with mutated AML1 (core) sites and found a few cases of second-site alterations. These consisted of deletions involving the NF1 sites and alterations in overall number of repeat elements, and they conferred increased enhancer strength in transient transcription assays. We have now tested the pathogenicity of a virus harboring one such second-site variant enhancer in inbred NMRI mice. It induced lymphomas with a 100% incidence and a significantly shorter latency than the AML1 mutant it evolved from. The enhancer structure thus represents the selection for a more tumorigenic virus variant during the pathogenic process. Sequencing of provirus from the induced tumors showed the new enhancer variant to be genetically stable. Also, Southern blotting showed that the tumors induced by the variant were T-cell lymphomas, as were the wild-type-induced lymphomas. In contrast, tumors induced by the original core/AML1 site I-II mutant appeared to be of non-T-cell origin and several proviral genomes with altered enhancer regions could be found in the tumors. Moreover, reporter constructs with the new tumor-derived variant could not be transactivated by AML1 in cotransfection experiments as could the wild type. These results emphasize the importance of both core/AML1 site I and site II for the pathogenic potential of SL3-3 and at the same time show that second-site alterations can form a viral variant with a substantial pathogenic potential although both AML1 sites I and II are nonfunctional. PMID: 9311802 [PubMed - indexed for MEDLINE] ---------------------------------------------------------------