IntroductionA growing body of evidence supports a role for members of the mammalian Hox homeobox gene family of transcription factors in the regulation of hematopoiesis. 1 In a subgroup of human myeloid leukemias, the HOXA9 gene is involved in a recurrent translocation between chromosomes 7 (NUP98) and 11 (HOXA9), resulting in a fusion transcript that encodes the NUP98-HOXA9 oncoprotein. 2,3 Recently, the expression of HOXA9 was also shown to be the single most highly correlating factor (of 6817 genes tested) for poor prognosis in human acute myeloid leukemia (AML), 4 possibly indicating a broader role for this gene in human leukemia beyond that caused by its chromosomal translocations.The leukemogenic potential of Hoxa9 was directly demonstrated by the development of AML in mouse bone marrow transplantation chimeras that received a graft of primitive hematopoietic cells engineered by retroviral gene transfer to overexpress Hoxa9. 5,6 Although the latency for the development of AML in these chimeras is much shorter than reported for other similar Hox gene chimeras (ie, Hoxa10, Hoxb3, Hoxb8), overexpression of Hoxa9 alone is not sufficient to induce leukemia, and additional somatic mutation(s) are needed. [6][7][8][9] In leukemic transformation, genetic interactions have been demonstrated between Hoxa9 and molecules such as Meis1 and E2A-PBX1, 5,6 both of which can bind DNA cooperatively with Hoxa9. 10,11 This suggests the presence of a leukemogenic complex, which includes Hoxa9 and potentially some of its DNA-binding cofactors.In normal human CD34 ϩ bone marrow cells, HOXA9 is preferentially expressed in subfractions that are highly enriched for primitive long-term culture-initiating cells or myeloid progenitor cells, and its expression is significantly reduced in the erythroid lineage. 12,13 This pattern of Hoxa9 expression is conserved in murine hematopoietic cells, with high levels detected in the Sca-1 ϩ lineage-negative (LinϪ) bone marrow subpopulation that is enriched for primitive hematopoietic cells, whereas much lower levels are present in the progenitor-depleted fraction (eg, Sca-1 Ϫ Lin ϩ ). 14,15 Hoxa9 is also expressed in purified subpopulations of B and T lymphoid cells, although at levels much lower than those found in primitive bone marrow cells. 14 Analysis of the Hoxa9 null mutant mice showed that lack of Hoxa9 caused a significant decrease in bone marrow myeloid and pre-B-cell progenitors, and Hoxa9 Ϫ/Ϫ myeloid progenitors have a blunted response to granulocyte colony-stimulating factor, a growth factor known to enhance the production and release of granulocytes from the bone marrow. 14 Although adult thymocyte development was only mildly abnormal in Hoxa9 Ϫ/Ϫ mice, the fetal thymus cellularity was reduced 5-to 10-fold, due in part to a delay in thymocyte progression from immature single-positive (T-cell receptor-negative) to double-positive (ie, CD4 ϩ CD8 ϩ ) T cells. 14,16 In contrast to lymphoid and myeloid progenitor cells in the Hoxa9 Ϫ/Ϫ mice, no significant decrease was detected in more...

Tumor-suppressor genes (TSGs) have been classically defined as genes whose loss of function in tumor cells contributes to the formation and/or maintenance of the tumor phenotype. TSGs containing nonsense mutations may not be expressed because of nonsense-mediated RNA decay (NMD). We combined inhibition of the NMD process, which clears transcripts that contain nonsense mutations, with the application of high-density single-nucleotide polymorphism arrays analysis to discriminate allelic content in order to identify candidate TSGs in five breast cancer cell lines. We identified ARID1A as a target of NMD in the T47D breast cancer cell line, likely as a consequence of a mutation in exon-9, which introduces a premature stop codon at position Q944. ARID1A encodes a human homolog of yeast SWI1, which is an integral member of the hSWI/SNF complex, an ATPdependent, chromatin-remodeling, multiple-subunit enzyme. Although we did not find any somatic mutations in 11 breast tumors, which show DNA copy-number loss at the 1p36 locus adjacent to ARID1A, we show that low ARID1A RNA or nuclear protein expression is associated with more aggressive breast cancer phenotypes, such as high tumor grade, in two independent cohorts of over 200 human breast cancer cases each. We also found that low ARID1A nuclear expression becomes more prevalent during the later stages of breast tumor progression. Finally, we found that ARID1A reexpression in the T47D cell line results in significant inhibition of colony formation in soft agar. These results suggest that ARID1A may be a candidate TSG in breast cancer.

cMutations in the gene encoding CLN5 are the cause of Finnish variant late infantile Neuronal Ceroid Lipofuscinosis (NCL), and the gene encoding CLN5 is 1 of 10 genes (encoding CLN1 to CLN9 and cathepsin D) whose germ line mutations result in a group of recessive disorders of childhood. Although CLN5 localizes to the lysosomal compartment, its function remains unknown. We have uncovered an interaction between CLN5 and sortilin, the lysosomal sorting receptor. However, CLN5, unlike prosaposin, does not require sortilin to localize to the lysosomal compartment. We demonstrate that in CLN5-depleted HeLa cells, the lysosomal sorting receptors sortilin and cation-independent mannose 6-phosphate receptor (CI-MPR) are degraded in lysosomes due to a defect in recruitment of the retromer (an endosome-to-Golgi compartment trafficking component). In addition, we show that the retromer recruitment machinery is also affected by CLN5 depletion, as we found less loaded Rab7, which is required to recruit retromer. Taken together, our results support a role for CLN5 in controlling the itinerary of the lysosomal sorting receptors by regulating retromer recruitment at the endosome.

The Hoxa9 and Meis1 genes represent important oncogenic collaborators activated in a significant proportion of human leukemias with genetic alterations in the MLL gene. In this study, we show that the transforming property of Meis1 is modulated by 3 conserved domains, namely the Pbx interaction motif (PIM), the homeodomain, and the C-terminal region recently described to possess transactivating properties. -HOXA9) that characterize myeloid leukemias in human and mice; (3) the overexpression of HOXA7 and HOXA9 in several MLL-induced human leukemias 11,12 ; (4) the distinctly high correlation between high levels of HOXA9 expression and poor prognosis in human acute myeloid leukemia (AML) 13 ; and (5) coexpression of BCR-ABL and NUP98-HOXA9 in a significant number of human leukemic specimens, 14 and a recent proof of their genetic interaction. 15 Together, these studies establish a direct and indirect role for multiple Hox genes in human leukemias, and highlight the importance of revealing the molecular bases of Hox-induced transformation.Some insight into the molecular mechanisms of Hox-induced transformation may be gained from studies of Hox cofactors, members of the TALE (for 3-amino-acid loop extension) family of homeodomain proteins. Of interest, founders of 2 subgroups within this family, namely PBX1 16,17 and Meis1, 18 were identified based on their participation in human and mouse leukemia, respectively.Hox, Pbx, and Meis participate in a multiprotein interaction network. The cooperative interaction between Pbx and Hox proteins enhances the DNA binding affinity and specificity of Hox proteins 19 and is essential for at least some of the Hox-dependent developmental programs. [20][21][22] In contrast, a functional role for a dimeric Hox-Meis complex has so far not been established. 23 Members of the Meis family can form complexes with Pbx in DNA-dependent and independent manners. [24][25][26] Interaction with Meis induces nuclear localization of Pbx by preventing its nuclear export 27,28 and promoting nuclear import. 29,30 Indirect interaction between Hox and Meis proteins was established by the identification of Hox-Pbx-Meis heterotrimeric complexes. 26 PBX1 is involved as part of the fusion E2a-PBX1 oncoprotein in a high proportion of human pre-B leukemias. 16,17 We recently generated a mouse model of E2a-PBX1-induced pre-B-cell leukemia, and showed that the Hoxa locus was targeted by murine Moloney leukemia virus (MMLV) in the majority of the leukemias analyzed. 31 These leukemias were characterized by aberrant expression of Hoxa genes, and all expressed high levels of Hoxa7, thus pointing to a genetic interaction between E2a-PBX1 and Hoxa genes. In support of these findings, we

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It is believed that hemopoietic stem cells (HSC),

The treatment of breast cancer has benefitted tremendously from the generation of estrogen receptor-a (ERa)-targeted therapies, but disease relapse continues to pose a challenge due to intrinsic or acquired drug resistance. In an effort to delineate potential predictive biomarkers of therapy responsiveness, multiple groups have identified several uncharacterized cofactors and interacting partners of ERa, including Split Ends (SPEN), a transcriptional corepressor. Here, we demonstrate a role for SPEN in ERa-expressing breast cancers. SPEN nonsense mutations were detectable in the ERa-expressing breast cancer cell line T47D and corresponded to undetectable protein levels. Further analysis of 101 primary breast tumors revealed that 23% displayed loss of heterozygosity at the SPEN locus and that 3% to 4% harbored somatically acquired mutations. A combination of in vitro and in vivo functional assays with microarray-based pathway analyses showed that SPEN functions as a tumor suppressor to regulate cell proliferation, tumor growth, and survival. We also found that SPEN binds ERa in a ligand-independent manner and negatively regulates the transcription of ERa targets. Moreover, we demonstrate that SPEN overexpression sensitizes hormone receptor-positive breast cancer cells to the apoptotic effects of tamoxifen, but has no effect on responsiveness to fulvestrant. Consistent with these findings, two independent datasets revealed that high SPEN protein and RNA expression in ERa-positive breast tumors predicted favorable outcome in patients treated with tamoxifen alone. Together, our data suggest that SPEN is a novel tumor-suppressor gene that may be clinically useful as a predictive biomarker of tamoxifen response in ERa-positive breast cancers. Cancer Res; 75(20); 4351-63. Ó2015 AACR.

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