Jay L. Hess

Disruptor of telomeric silencing 1-like (Dot1l) is a histone 3 lysine 79 methyltransferase. Studies of constitutive Dot1l knockout mice show that Dot1l is essential for embryonic development and prenatal hematopoiesis. DOT1L also interacts with translocation partners of Mixed Lineage Leukemia (MLL) gene, which is commonly translocated in human leukemia. However, the requirement of Dot1l in postnatal hematopoiesis and leukemogenesis of MLL translocation proteins has not been conclusively shown. With a conditional Dot1l knockout mouse model, we examined the consequences of Dot1l loss in postnatal hematopoiesis and MLL translocation leukemia. Deletion of Dot1l led to pancytopenia and failure of hematopoietic homeostasis, and Dot1l-deficient cells minimally reconstituted recipient bone marrow in competitive transplantation experiments. In addition, MLL-AF9 cells required Dot1l for oncogenic transformation, whereas cells with other leukemic oncogenes, such as Hoxa9/Meis1 and E2A-HLF, did not. These findings illustrate a crucial role of Dot1l in normal hematopoiesis and leukemogenesis of specific oncogenes.

Received: September 8, 2010, Accepted: October 5, 2010, Published: October 5, 2010 // <![CDATA[ $('#abstractHeader').hide(); // ]]> Recent studies identified an interaction between the Polymerase Associated Factor complex (PAFc) and Mixed Lineage Leukemia (MLL), including MLL-rearranged oncoproteins. This interaction is critical for MLL transcriptional activity and MLL-rearranged leukemogenesis. Here, we discuss the potential molecular role of the PAFc in transcriptional dysregulation of MLL target genes and the interplay between PAFc and MLL-rearranged oncoproteins in leukemogenesis.

Disruption of epigenetic regulators of transcription is a central mechanism of oncogenesis. Many of the advances in the understanding of these mechanisms are attributable to the successful development of chromatin immunoprecipitation (ChIP) for in vivo detection of histone modifications as well as chromatin binding regulatory proteins. This is a powerful technique for analyzing histone modifications as well as binding sites for proteins that bind either directly or indirectly to DNA. Here we present two ChIP protocols. The first is particularly useful for identifying histone modifications or binding at specific, known genomic sites. The second, employing serial analysis of gene expression, is particularly powerful for the discovery of previously unidentified sites of modification or binding.

In the hematopoietic system, menin was found to interact with MLL, a large protein encoded by the mixed linage leukemia gene that acts as a histone H3 methyltransferase. The MLL gene is a recurrent target for translocations in both acute myeloid and acute lymphoid leukemias. MLL gene rearrangements involve a variety of translocation partners, giving rise to MLL fusion proteins whose transforming ability is mediated through upregulated expression of Homeobox (Hox) genes as well as other targets. Recent work indicates that menin is an essential partner of MLL fusion proteins in leukemic cells and that it regulates normal hematopoiesis. In the absence of menin, steady-state hematopoiesis is largely preserved; however, menin-deficient hematopoietic stem cells are markedly deficient in situations of hematopoietic stress, such as during recovery after bone marrow transplantation. In leukemias driven by MLL fusion proteins, menin is essential for transformation and growth of the malignant cells. Thus, menin-MLL interactions represent a promising therapeutic target in leukemias with MLL rearrangements.

HOX and three amino acid loop extension (TALE) proteins cooperate to induce transformation in mouse leukemia models, and are dysregulated in a variety of human leukemias. Despite decades of research, the mechanism of action for Hox proteins in embryogenesis and hematopoiesis remains unclear. Recent studies on the roles of Hoxa9 and Meis1 in leukemia has led to a wealth of new data, but their molecular mechanisms of action and synergy remain obscure. Advances in genome-wide technologies offer new avenues for understanding how homeodomain-containing transcription factors exert their programs in normal and neoplastic development.

Fusion of the mixed lineage leukemia protein (MLL) to one of over 50 different translocation partners converts it into a potent leukemogenic oncoprotein. The resulting fusion proteins transform primarily through upregulation of A-cluster Hox genes, including Hoxa9 and the Hox cofactor Meis1. Considerable progress has been made in delineating the differences between normal Hox gene regulation by MLL and deregulated transcription in MLL-induced leukemias. Some MLL translocation partners dimerize the truncated MLL molecule. Other translocation partners appear to recruit nuclear coactivator complexes that have diverse enzymatic activities that impinge on transcriptional elongation pathways. These enzymatic activities, including RNA polymerase II phosphorylation as well as histone H3 lysine 79 methylation present attractive targets for the development of future MLL-directed therapy.