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 in IntroductionThe disruption of the human homologue of the Drosophila Trithorax (trx) gene, MLL on 11q23 1-4 by chromosomal translocations is a frequent event in human acute leukemia. These translocations, leading to the juxtaposition of genetic elements and formation of MLL fusion genes, occur in approximately 5% to 10% of human acute leukemias, but with a higher frequency in infant leukemias 5 and secondary leukemias. 6 Although the full biological function of MLL is uncertain, it is known to act as a positive regulator of HOX gene expression in development. 7,8 Currently, 20 different translocations affecting the MLL gene have been molecularly cloned and the partner genes identified. 9 Significantly, all such translocations result in in-frame fusions at the messenger RNA level, and, therefore, the rearrangements result in the production of chimeric proteins in which the N-terminus of MLL is consistently fused to the C-terminus encoded by the partner gene. [10][11][12] Because such a diverse group of proteins can be fused to MLL, the role of the fusion proteins in leukemogenesis remains obscure. 13 Although most of the fusion partners are structurally and functionally unrelated to each other, 14 a number are involved in transcriptional regulation. For example, ENL, AF9, and AF4 activate transcription from synthetic reporter genes in vivo. [15][16][17][18][19][20] The AF10 gene is one of the few MLL partner genes to be independently rearranged with a third gene in leukemia, the CALM gene in the t(10;11)(p12;q14) translocation. 21 AF10 complementary DNA (cDNA) encodes a 1084-aa protein, a member of a family of proteins including MLL, all carrying a conserved LAP/PHD finger domain. 22,23 There is also a putative AT-hook motif, 24 a bipartite nuclear localization signal, a leucine zipper domain, and a glutamine-rich region at the C-terminus. The latter is not present in all isoforms, as a result of alternative splicing. 25,26 Although different breakpoints have been described for AF10, the resultant fusion protein in both products consistently loses the...

As a result of the synovial sarcoma associated t(X;18) translocation, the human SYT gene on chromosome 18 is fused to either the SSX1 or the SSX2 gene on the X chromosome. Although preliminary evidence indicates that the (fusion) proteins encoded by these genes may play a role in transcriptional regulation, little is known about their exact function. We set out to isolate interacting proteins through yeast two hybrid screening of a human cDNA library using SYT as a bait. Of the positive clones isolated, two were found to correspond to the acute leukemia t(10;11) associated AF10 gene, a fusion partner of MLL. Con®rmation of these results was obtained via co-immunoprecipitation of endogenous and exogenous, epitope-tagged, SYT and AF10 proteins from cell line extracts and colocalization of epitopetagged SYT and AF10 proteins in transfected cells. Subsequent sequential mutation analysis revealed a highly speci®c interaction of N-terminal SYT fragments with C-terminal AF10 fragments. The N-terminal interaction domain of the SYT protein was also found to be present in several SYT orthologs and homologs. The C-terminal interaction domain of AF10 is located outside known functional domains. Based on these results, a model is proposed in which the SYT and AF10 proteins act in concert as bipartite transcription factors. This model has implications for the molecular mechanisms underlying the development of both human synovial sarcomas and acute leukemias. Oncogene (2001) 20, 3281 ± 3289.

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