👤 Tracy Chaplin

The AF10 gene encodes a putative transcription factor containing an N-terminal LAP/PHD zinc finger motif, a functional nuclear localization signal, an AT-hook domain, and a leucine zipper toward the C-terminus. AF10 is involved in 2 distinct chromosomal translocations associated with hematologic malignancy. The chimeric fusion proteins MLL/AF10 and CALM/AF10, resulting from the t(10;11)(p12;q23) and the t(10;11)(p12;q14), respectively, consistently retain the leucine zipper motif of AF10. This part of the C-terminal region was used as bait in a yeast 2 hybrid screening of a testis complementary DNA library. The leucine zipper interacted with GAS41, a protein previously identified as the product of an amplified gene in a glioblastoma. GAS41 shows significant homology to the Saccharomyces cerevisiae protein ANC1 and to the human MLL fusion partners AF9 and ENL. The interaction was confirmed in vivo. Furthermore, the study showed by coimmunoprecipitation that GAS41 interacts with INI1 (Integrase Interactor 1) and that INI1 was present in the AF10 immunoprecipitate. INI1 is the human homologue of the yeast SNF5 protein, a component of the SWI/SNF complex, which acts to remodel chromatin and to modulate transcription. The retention of the leucine zipper in the MLL and CALM fusions suggests that a key feature of these chimeric proteins may be their ability to interfere in normal gene regulation through interaction with the adenosine triphosphate-dependent chromatinremodeling complexes. Show less

The cell line U937, which has been used extensively for studies of myeloid differentiation, bears the t(10;11)(p13;q14) translocation which results in a fusion between the MLLT10 (myeloid/lymphoid or mixed-lineage leukemia [trithorax, Drosophila, homolog]; translocated to 10; alias AF10) gene and the Ap-3-like clathrin assembly protein, PICALM (Clathrin assembly lymphoid myeloid leukaemia). Apart from this translocation, very little is known about the other genetic alterations in this cell line that may represent significant events in disease progression. In this study, conventional G-banding, CGH and M-FISH have been used to characterise fully all of the cytogenetic alterations present in the U937 cell line. M-FISH analysis confirmed the presence of the t(10;11) and an apparently normal copy of both chromosomes 10 and 11. A t(1;5) translocation was observed as well as several unbalanced rearrangements. CGH detected amplifications resulting from duplications of 2q, 6p and 13q. These changes could result in fusion gene products involved in carcinogenesis or the positions of putative oncogenes and tumour suppressor genes. A good correlation between conventional G-banding, CGH and M-FISH was observed. Show less

The interstitial insertion of genetic material from one chromosome into another can achieve the type of gene-gene fusions more usually associated with chromosome translocations. An example of such an interstitial insertion, which has created an MLL-AF10 fusion in an acute myeloid leukaemia, has been analysed at the genomic level. The genomic fusion, which resulted in the juxtaposition of 3' AF10 sequence to 5' MLL sequence, was identified within MLL and AF10 intronic sequences. It was further established that the remaining 3' MLL sequence, from exon 6 onwards, was fused to novel sequence of unknown origin (named FM3 for fused to MLL 3'). The points of fusion of these 5' and 3' portions of MLL matched to adjacent nucleotides and lay between exons 5 and 6. The FM3 sequence was shown to be from chromosome arm 10p and located close to AF10 in a proximal position. It was subsequently demonstrated that in the leukaemia a third fusion existed between 5' AF10 and the FM3 sequence at a point immediately downstream from its fusion to MLL. It was therefore concluded that the MLL-AF10 gene fusion is the result of a simultaneous transposition of genetic material into the MLL gene and the joining of the remaining free ends on chromosome 10. This kind of event, characterised completely here for the first time, is a means to achieve a fusion when the genes involved lie in opposite orientations and results in three genomic junctions. Show less

The MLL gene is reciprocally translocated with one of a number of different partner genes in a proportion of human acute leukaemias. The precise mechanism of oncogenic transformation is unclear since most of the partner genes encode unrelated proteins. However, two partner genes, AF10 and AF17 are related through the presence of a cysteine rich region and a leucine zipper. The identification of other proteins with these structures will aid our understanding of their role in normal and leukaemic cells. We report the cloning of a novel human gene (BRL) which encodes a protein containing a cysteine rich region related to that of AF10 and AF17 and is overall most closely related to the previously known protein BR140. BRL maps to chromosome 22q13 and shows high levels of expression in testis and several cell lines. The deduced protein sequence also contains a bromodomain, four potential LXXLL motifs and four predicted nuclear localization signals. A monoclonal antibody raised to a BRL peptide sequence confirmed its widespread expression as a 120 Kd protein and demonstrated localization to the nucleus within spermatocytes. Show less

We have cloned Af10, the murine homologue of the MLL partner gene AF10. The predicted open reading frame of Af10 contains 1069 aa which are 90% identical to those of AF10. Af10 contains an N-terminal cysteine-rich region with a LAP/PHD finger, a leucine zipper domain and a glutamine-rich region at the C-terminus, features also found in the human proteins AF10 and AF17. A single 5. 5-kb transcript was detected in murine tissues with the highest level of expression in the testes. A polyclonal antibody raised to the cysteine-rich region of AF10 was able to identify a double band of 140 kDa on Western analysis in mouse testicular extracts. After subcellular separation Af10 was identified in both the nuclear and cytoplasmic extracts, again as a double band of 140 kDa in size. In situ hybridisation studies were performed with sense and antisense digoxigenin-labelled oligonucleotides. High levels of expression were noted in postmeiotic germ cells, especially in spermatids from around stage VI to stage VIII. High levels of expression were also seen in the white matter of the cerebellum, extending into the granular layer. The expression in differentiated rather than in proliferating cells suggests that the role of Af10 may lie in the suppression of proliferation rather than in differentiation. Since the LAP/PHD finger domains are lost in the MLL-AF10 fusion, arguably such a function could be carried out by this domain. Show less

We have identified and further characterized a Caenorhabditis elegans gene, CEZF, that encodes a protein with substantial homology to the zinc finger and leucine zipper motifs of the human gene products AF10, MLLT6, and BR140. The first part of the zinc finger region of CEZF has strong similarity to the corresponding regions of AF10 (66%) and MLLT6 (64%) at the cDNA level. As this region is structurally different from previously described zinc finger motifs, sequence homology searches were done. Twenty-five other proteins with a similar motif were identified. Because the functional domain of this motif is potentially disrupted in leukemia-associated chromosomal translocations, we propose the name of leukemia-associated protein (LAP) finger. On the basis of these comparisons, the LAP domain consensus sequence is Cys1-Xaa1-2-Cys2-Xaa9-21-Cys3-Xaa2-4 -Cys4-Xaa4-5-His5-Xaa2-Cys6-Xaa12-46 - Cys7-Xaa2-Cys8, where subscripted numbers represent the number of amino acid residues. We review the evidence that this motif binds zinc, is the important DNA-binding domain in this group of regulatory proteins, and may be involved in leukemogenesis. Show less

The gene on chromosome 10 at band p12 (AF10), involved in the t(10;11) translocation in acute myeloid leukemia, has been identified and shown to contain conserved zinc finger and leucine zipper domains. These regions are highly homologous to the equivalent regions on AF17, the gene involved in the t(11;17) translocations. A series of adult, childhood, and infant leukemias with either simple or complex versions of the t(10;11) has been examined by Southern analysis and shown to involve rearrangement to the HRX locus. Reverse transcriptase-polymerase chain reaction from either bone marrow or peripheral blood cells showed that HRX sequence was fused to AF10 sequence in all 8 cases and subsequent sequence analysis showed an in-frame fusion between the HRX and AF10 sequence. A consistent feature of these fusions was the juxtaposition of the leucine dimerization motif of AF10 onto the NH2-terminal region of HRX. The published data suggest that a similar conclusion can be drawn about the t(11;17) translocation, implying a critical role for this motif in the chimaeric HRX protein. Show less

A novel class of conserved transcription factors has been identified from the molecular cloning of AF10, the gene involved in the t(10;11)(p12;q23) translocation of acute myeloid leukemias. AF10 encodes a 109-kD protein of 1,027 amino acids and contains an N-terminal zinc finger region and a C-terminal leucine zipper. These structures have been found to be conserved in sequence and position in three other proteins, AF17, BR140, and a previously unrecognized Caenorhabditis elegans gene, provisionally named CEZF. The overall structure, level of sequence conservation, and expression pattern suggest that these genes encode a new class of transcription factors, some of which are targets for chromosomal translocation in acute leukemia. Show less

A 280 kilobase (kb) contig was isolated from mouse genomic P1 and cosmid libraries, using as probes human cDNA and genomic DNA fragments that map in the interval between the second component of complement and tumor necrosis factor genes of the HLA complex. The clone contig demonstrates synteny of eleven mouse genes that are homologous to genes initially mapped within the human major histocompatibility complex. These include the mouse homologs of BAT2 (HLA-B-associated transcript 2) through BAT9 and also three HSP70-related genes. Five P1 clones form a contig of 240 kb that spans from BAT9 through BAT3. Twelve cosmid clones are arranged in three contigs that confirm most of the structure of the P1 contig and link the mouse BAT3 homolog to the BAT2 homolog approximately 15 kb farther telomeric. Polymorphic DNA markers within the cloned region were used to map the cleft palate susceptibility-1 (Cps-1) locus to the interval between Hsp70.1 and BAT6 (valyl-tRNA synthetase). This refines the location of the Cps-1 locus to a 45 kb region contained in the H2-124 P1 insert. Show less