Mouse Af9 Is a Controller of Embryo Patterning, Like Mll, Whose Human Homologue Fuses with AF9 after Chromosomal Translocation in Leukemia

Abstract

Chromosomal translocation t(9;11)(p22;q23) in acute myeloid leukemia fuses the MLL and AF9 genes. We have inactivated the murine homologue of AF9 to elucidate its normal role. No effect on hematopoiesis was observed in mice with a null mutation of Af9. However, an Af9 null mutation caused perinatal lethality, and homozygous mice exhibited anomalies of the axial skeleton. Both the cervical and thoracic regions were affected by anterior homeotic transformation. Strikingly, mice lacking functional Af9 exhibited a grossly deformed atlas and an extra cervical vertebra. To determine the molecular mediators of this phenotype, analysis of Hox gene expression by in situ hybridization showed that Af9 null embryos have posterior changes in Hoxd4 gene expression. We conclude that the Af9 gene is required for normal embryogenesis in mice by controlling pattern formation, apparently via control of Hox gene regulation. This is analogous to the role of Mll, the murine homolog of human MLL, to which the Af9 gene fuses in acute myeloid leukemias.The consequence of chromosomal translocations in human cancer is to cause enforced expression of proto-oncogenes or the creation of tumor-specific fusion genes (reviewed in reference 52). Chromosomal translocations often involve transcription regulators (9) which function as master regulators of cell fate in their normal situations of expression (53). Human AF9 was identified as one of the most common fusion partners of the mixed-lineage leukemia protein (MLL, also called HTRX and ALL-1) (14,20,63,70) and is most usually associated with acute myeloid leukemias (AML). MLL fusion proteins are present in approximately 10% of acute lymphoid leukemias and myeloid leukemias (42) and in up to 80% of leukemias in infants (33, 51), as well as in 85% of cases of secondary leukemias developing after treatment of a primary tumor with alkylating agents or topoisomerase II inhibitors (3,8,61). In an individual leukemia, the fusion protein results from one of a variety of reciprocal chromosomal translocations, causing the association of upstream exons of the MLL gene on chromosome 11, band q23, with downstream exons of the partner gene, generally on a separate chromosome. MLL is a human homologue of Drosophila melanogaster trithorax (trx), a master homeotic gene regulator essential for normal patterning during embryo development (2, 43). Gene-targeting studies have demonstrated a similar role for MLL in mammals (17,21,67,69).More than 30 different MLL fusions have been identified (28), and these code for a structurally and functionally heterogeneous group of proteins (reviewed in references 1 and 10). However, different MLL fusion proteins are consistently associated with hematopoietic tumors of different lineages. Thus, whereas MLL-AF9 is mainly found in AML, the translocation involving the AF4 gene (16,20,47) occurs almost exclusively in tumors of the B-cell lineage. This suggests that the fusion partner plays a role in determining disease phenotype, either because it influences the site and timing of...

Characterization of Bcor expression in mouse development

“…RNAi knock-down of Bcor in zebrafish results in eye, skeletal and nervous system abnormalities consistent with those found in MCOPS2 patients (Ng et al, 2004). BCOR has also been shown to interact with and repress transcriptional activation of the MLL fusion protein AF9 (Srinivasan et al, 2003), a known regulator of Hox gene expression (Collins et al, 2002) and skeletal development, further implicating BCOR as a key developmental regulator. The pleiotropic effects induced by the loss of functional BCOR in humans and zebrafish (Ng et al, 2004) clearly illustrate the essential role of BCOR during embryogenesis and emphasizes the importance of determining the spatial and temporal expression of BCOR during development.…”

Proteomics Analysis of Ring1B/Rnf2 Interactors Identifies a Novel Complex with the Fbxl10/Jhdm1B Histone Demethylase and the Bcl6 Interacting Corepressor

“…Sequence coverage and peptide numbers for these proteins as shown in Table I would indicate that the Ring1B/Rnf2-Pc3/Cbx8-ENL/Mllt1 may form a multiprotein complex of lower abundance and may also contain additional, as yet uncharacterized components that are very likely included in Table I. A clue as to the functional activity of these complexes may be found in the phenotypes of Af9/ Mllt3-deficient mice, namely the anteriorization of the rostral boundary of expression of a Hox gene and the alterations of the axial skeleton (62), which are reminiscent of those seen in PcG mutants.…”

The mixed lineage leukemia fusion partner AF9 binds specific isoforms of the BCL-6 corepressor

“…However, the function of these proteins in normal and diseased cells is not yet clear. Targeted disruption of the AF9 and ENL loci in mice has recently been accomplished and the lethal phenotype of nullizygous embryos (ENL) or newborn pups (AF9) indicates that each gene plays an essential role in embryonic development (Collins et al, 2002;Doty et al, 2002). Artificial reporter gene assays indicate that the Ctermini of the proteins promote transcription but, apart from these assays, there is little direct evidence that AF9 and ENL are bona fide transcription factors (Rubnitz et al, 1994;this report).…”