In C. elegans embryos, a Wnt/MAPK signaling pathway downregulates the TCF/LEF transcription factor POP-1, resulting in a lower nuclear level in signal-responsive cells compared to their sisters. Although the beta-catenin WRM-1 is required for POP-1 downregulation, a direct interaction between these two proteins does not seem to be required, as the beta-catenin-interacting domain of POP-1 is dispensable for both POP-1 downregulation and function in early embryos. We show here that WRM-1 downregulates POP-1 by promoting its phosphorylation by the MAP kinase LIT-1 and subsequent nuclear export via a 14-3-3 protein, PAR-5. In signal-responsive cells, we also detect a concurrent upregulation of nuclear LIT-1 that is dependent on Wnt/MAPK signaling. Our results suggest a model whereby Wnt/MAPK signaling downregulates POP-1 levels in responsive cells, in part by increasing nuclear LIT-1 levels, thereby increasing POP-1 phosphorylation and PAR-5-mediated nuclear export.

IntroductionAcute myeloid leukemia (AML) is a heterogeneous disease that is classified based on the presence of specific cytogenetic abnormalities as well as the French-American-British (FAB) classification of the leukemic cells and immunophenotype. One of the common translocations identified in leukemia is between chromosome 8q22 and chromosome 21q22 ( Figure 1a). 1 It is associated with nearly 40% of cases of FAB-M2 AML and 8% to 20% of all cases of AML depending on the genetic background and geographic location of the population. The (8;21) translocation is also observed in approximately 6% of AML M1 and, more rarely, in AML M0, M4, M5, and other myeloproliferative syndromes. 2,3 The involved genes are, on chromosome 8, MTG8 or ETO, meaning myeloid translocation gene or eight twenty-one, respectively, 4,5 and AML1 (acute myeloid leukemia factor 1) on chromosome 21. 4 The commonly used name for the t(8;21) fusion protein is AML1-MTG8 or AML1-ETO, and we refer to it as AML1-ETO in this review. AML1 was also discovered from other studies that are not related to t(8;21) and has several different names. 6 Its HUGO (Nomenclature Committee of the Human Genome Organization) name is RUNX1. In correlation, MTG8/ETO is named RUNX1T1 for RUNX1 translocation 1.The t(8;21) generates the 2 fusion genes AML1-ETO and ETO-AML1 ( Figure 1B). AML1-ETO mRNA is easily detectable using polymerase chain reaction (PCR) primers on 2 sides of the fusion point. However, ETO-AML1 mRNA was not identified using a similar approach (E. Kanbe, D.-E.Z., unpublished data, February 2003). This result indicates that the ETO-AML1 transcript is not expressed, is expressed at an extremely low level, or is highly unstable due to degradation. All of the studies on t(8;21) have therefore focused on AML1-ETO.Most of the coding region of the ETO gene is fused to the AML1 amino terminus containing the DNA-binding runt homology domain (RHD) to generate an AML1-ETO fusion protein ( Figure 1C). 4,5,7 The ETO gene has 14 exons. The original cloned AML1-ETO cDNA contained ETO exons 2 through 11; the fusion transcript produces an AML1-ETO protein of 752 amino acids ( Figure 1C). 8 The ETO portion of the full-length AML1-ETO protein contains 3 proline-serine-threonine (PST)-rich regions and 4 Nervy homology regions (NHR1-4) ( Figure 1C). 9 The PST-rich regions have multiple potential kinase phosphorylation sites (SP [Serine-Proline] and TP [Threonine-Proline]). Phosphorylation of ETO has been reported although no kinase involved in its phosphorylation has been identified. 10 NHR1, also called the TAF (TATA box binding protein associated factor) homology domain, shares a sequence similarity with TAF110 and other related TAFs. NHR2 has a hydrophobic amino acid heptad repeat, which is critical for ETO oligomerization. 11 NHR3 contains a predicted coiled-coil structure. NHR4 is a myeloid-Nervy-DEAF1 (MYND) homology domain with 2 predicted zinc finger motifs.Expression of the AML1-ETO fusion gene is under the control of the AML1 promoter. The AML1 gene has 2 promot...

The transcription factor BCL11A has recently been reported to be a driving force in triple-negative breast cancer (TNBC), contributing to the maintenance of a chemoresistant breast cancer stem cell (BCSC) population. Although BCL11A was shown to suppress γ-globin and p21 and to induce MDM2 expression in the hematopoietic system, its downstream targets in TNBC are still unclear. For its role in transcriptional repression, BCL11A was found to interact with several corepressor complexes; however, the mechanisms underlying these interactions remain unknown. Here, we reveal that BCL11A interacts with histone methyltransferase (PRC2) and histone deacetylase (NuRD and SIN3A) complexes through their common subunit, RBBP4/7. In fluorescence polarization assays, we show that BCL11A competes with histone H3 for binding to the negatively charged top face of RBBP4. To define that interaction, we solved the crystal structure of RBBP4 in complex with an N-terminal peptide of BCL11A (residues 2-16, BCL11A(2-16)). The crystal structure identifies novel interactions between BCL11A and the side of the β-propeller of RBBP4 that are not seen with histone H3. We next show that BCL11A(2-16) pulls down RBBP4, RBBP7, and other components of PRC2, NuRD, and SIN3A from the cell lysate of the TNBC cell line SUM149. Furthermore, we demonstrate the therapeutic potential of targeting the RBBP4-BCL11A binding by showing that a BCL11A peptide can decrease aldehyde dehydrogenase-positive BCSCs and mammosphere formation capacity in SUM149. Together, our findings have uncovered a previously unidentified mechanism that BCL11A may use to recruit epigenetic complexes to regulate transcription and promote tumorigenesis.

IntroductionAcute myeloid leukemia (AML) is a common hematologic malignancy characterized by an abnormal accumulation of myeloid precursors in the bone marrow and blood. Similar to many other types of cancer, genetic abnormalities are associated with the development of AML, particularly chromosomal translocations that result in novel fusion proteins. One of the common translocations implicated in AML is the 8q22;21q22 translocation [t(8;21)]. 1 Based on the French-American-British (FAB) classification of leukemic cells, t(8;21) is associated with nearly 40% of the AML cases with the FAB M2 phenotype. 2 t(8,21) involves the AML1 (RUNX1) gene on chromosome 21 and the ETO (MTG8, RUNX1T1) gene on chromosome 8. [3][4][5] AML1 is the DNA-binding subunit of the core binding factor (CBF) transcription factor complex. Its N-terminus contains a highly conserved DNA binding domain called the runt homology domain (RHD). t(8;21) fuses the N-terminus of AML1 including RHD in-frame with almost the entire ETO protein to form AML1-ETO. [3][4][5][6] This fusion protein acts as a dominant negative form of AML1 during embryogenesis. 7,8 It functions as a transcriptional repressor by interacting with NCoR/SMRT/HDAC. 9,10 AML1-ETO was shown to activate expression of BCL-2 and p21, possibly via interacting with p300. [11][12][13] AML1-ETO promotes stem cell renewal and blocks hematopoietic differentiation. [14][15][16] However, its role in blocking cell-cycle progression and promoting apoptosis contradicts its function in promoting leukemogenesis and therefore requires secondary mutagenic events for full transformation. 17,18 We previously identified a single nucleotide insertion that resulted in a truncated AML1-ETO protein (AML1-ETOtr or AEtr), which rapidly promoted leukemia. 19 Subsequently, we identified a C-terminally truncated variant of AML1-ETO named AML1-ETO9a (AE9a), resulting from alternative splicing and found to coexist with full-length AML1-ETO in most analyzed t(8;21) AML patients. 20 Similar to AEtr, AE9a causes a rapid onset of leukemia in mice, 20 which provides a useful mouse model to study the molecular biology of t(8;21) leukemia.To understand the molecular mechanism of AML1-ETOrelated leukemia development and to explore novel therapeutic targets to treat this type of leukemia, in this study we combined gene expression microarray and promoter occupancy (ChIP-chip) analyses to identify genes directly modulated by AE9a in primary murine leukemia cells. Among the common targets of microarray and ChIP-chip assays, approximately 30% show human t(8;21)-specific up or down-regulation compared with AML that have other chromosomal abnormalities. CD45, a protein tyrosine phosphatase and a negative regulator of cytokine/growth factor receptor and JAK/STAT signaling, 21-23 is among those targets. Its expression is down-regulated in AE9a leukemia cells. Consequently, JAK/STAT signaling is enhanced in these leukemia cells. We show that re-expression of CD45 suppresses JAK/STAT activation, delays leukemia development by AE9a an...

Triple-negative breast cancer (TNBC) is the most difficult subtype of breast cancer to treat due to a paucity of effective targeted therapies. Many studies have reported that breast cancer stem cells (BCSCs) are enriched in TNBC and are responsible for chemoresistance and metastasis. In this study, we identify LRP8 as a novel positive regulator of BCSCs in TNBC. LRP8 is highly expressed in TNBC compared to other breast cancer subtypes and its genomic locus is amplified in 24% of TNBC tumors. Knockdown of LRP8 in TNBC cell lines inhibits Wnt/β-catenin signaling, decreases BCSCs, and suppresses tumorigenic potential in xenograft models. LRP8 knockdown also induces a more differentiated, luminal-epithelial phenotype and thus sensitizes the TNBC cells to chemotherapy. Together, our study highlights LRP8 as a novel therapeutic target for TNBC as inhibition of LRP8 can attenuate Wnt/β-catenin signaling to suppress BCSCs.