MINI REVIEW New Insight Into the Molecular Mechanisms of MLL

MINI REVIEW

New insight into the molecular mechanisms of MLL-associated leukemia

Z-Y Li1, D-P Liu1 and C-C Liang1

1National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, 5 Dong Dan San Tiao, Beijing, PR China

Rearrangements of the MLL gene (ALL1, HRX, and Hrtx) located have been described, and 33 of the presumptive gene partners of at chromosome band 11q23 are commonly involved in adult MLL cloned and analyzed at the molecular level.13 The and pediatric cases of primary acute leukemias and also found in cases of therapy-related secondary leukemias. Studies on mechanisms by which these rearrangements result in leukemia mouse models of MLL translocation and cell lines containing remain largely unknown. MLL rearrangements showed that the MLL gene linked In addition to chromosomal translocations, other mechanisms chromosomal rearrangements to cellular differentiation and of MLL rearrangement have been demonstrated in patients with tumor tropism. Moreover, recent structural/functional studies acute leukemia. The partial tandem duplication (PTD) of MLL is on MLL and aberrant MLL proteins provided new clues and present in approximately 10% of patients with acute myeloid suggested that different mechanisms might be included in 14 leukemogenesis by MLL rearrangements. The connection leukemia (AML) and normal cytogenetics and in the majority between these different mechanisms will help us understand of patients with AML and trisomy 11 as the sole cytogenetic 15 globally how aberrant MLL oncogenes affect the normal cellular abnormality. This rearrangement is characterized by an processes at molecular level. internal duplication of MLL spanning exons 2–6 or 2–8.16 Leukemia (2005) 19, 183–190. doi:10.1038/sj.leu.2403602 Ampliﬁcation within chromosome arm 11q involving MLL gene Published online 16 December 2004 locus is a rare but recurrent aberration in AML and myelodys- Keywords: ; leukemia; molecular mechanisms MLL plastic syndrome (AML/MDS).17–19 These alternative classes of MLL mutants presumably promote leukemogenesis without the concomitant requirement for fusion of MLL to a partner protein, suggesting that alteration of MLL alone, in some circumstances, Introduction is sufﬁcient to promote leukemogenesis.

Leukemia is a heterogeneous disease at the molecular level resulting from a variety of alterations in numerous genes Functions of normal MLL protein important for cell growth, differentiation, and cell death. Identification and characterization of these genetic rearrange- MLL, a human homologue of the epigenetic transcriptional ments have been proved invaluable for appropriate diagnosis regulator Trithorax of Drosophila,20 is an upstream transcrip- and prognosis, especially in acute leukemia. Rearrangements of tional effector of HOX genes.21,22 The importance of normal the MLL gene (ALL1, HRX, and Hrtx) located at chromosome MLL protein for normal axial-skeletal developmental process band 11q23 are commonly involved in acute leukemia. These and HOX gene regulation has been demonstrated in the rearrangements have been shown to be associated with some embryos of heterozygous and homozygous MLL knockout21 1–3 4–6 adult and pediatric cases of primary acute leukemias and and MLL truncation mutant mice.23 Furthermore, expression of also are found in the majority of patients with secondary MLL protein is not necessary for turning on transcription of leukemias after prior treatment with DNA topoisomerase II certain HOX genes, but for the maintenance of their transcrip- 7,8 inhibitors (eg, etoposide). Unlike many other types of tion.22 Experiments in vitro using hematopoietic progenitors leukemia, the presence of an MLL translocation predicts early from embryos of homozygous MLL knockout mice24,25 or mice 9,10 relapse and poor prognosis. Frequently, leukemic blasts are with MLL mutant26 showed that MLL was also critical for characterized by the expression of lymphoid and myeloid hematopoietic development. Recent findings suggested that surface markers, defining MLL or Mixed Lineage Leukemia MLL is required during embryogenesis for the specification or 11 gene. expansion of hematopoietic stem cells.27 As HOX genes also The Mixed-Lineage Leukemia gene (MLL/HRX/ALL1) consists play a key role in the regulation of hematopoietic develop- of at least 36 exons, encoding a 3969 amino-acid nuclear ment,28 the hematopoietic dysfunction of MLL null cells is likely protein with a molecular weight of nearly 430 kDa that is to be attributed to deregulated patterns of HOX gene expression thought to function as a positive regulator of gene expression in in hematopoietic stem cells or progenitors. This link between early embryonic development and hematopoiesis. MLL translo- MLL, HOX gene regulation, and hematopoiesis is of particular cation breakpoints cluster within an 8.3-kb region spanning importance. 12 exons 5–11. To date, over 60 chromosome partners of 11q23 Structural/functional studies on normal MLL protein also provide some information on MLL functions. MLL protein Correspondence: Dr D-P Liu, National Laboratory of Medical possesses some functional domains that could confer significant Molecular Biology, Institute of Basic Medical Sciences, Chinese functional activity (Figure 1). Of these functional domains, AT Academy of Medical Sciences and Peking Union Medical College, 5 hooks and MT domain have DNA-binding activity that could be Dong Dan San Tiao, Beijing 100005, PR China; Fax: þ 8610 65105093; E-mail: [email protected] involved in the regulation of gene transcription through direct Received 22 July 2004; accepted 15 October 2004; Published online binding to DNA. The AT hook, with homology to the high- 16 December 2004 mobility group I (HMG-I) protein, binds to AT-rich regions of the Molecular mechanisms of MLL-associated leukemia Z-Y Li et al 184

Figure 1 Schematic depictions of wild type and aberrant MLL proteins. As MLL translocation occurs, MLL N terminus fuses with other nonrelated proteins, and they will form chimeric proteins. However, in the normal process, MLL will be cleaved into two fragments. The partial tandem duplication of MLL leads to fusion of the 50 portion of MLL protein with itself. Various putative functional domains are shown as colored boxes and labeled as follows: AT hooks, AT hook DNA binding motifs; SNL 1 and 2, speckled nuclear localization signals 1 and 2; CxxC (MT), cysteine-rich motif homologous to DNA methyltransferase and MBD1; PHD 1–3, PHD ﬁngers 1, 2 and 3; BROMO, BROMO domain; FYRN, F/Y- rich region, N-terminus; FYRC, F/Y-rich region, C-terminus; TAD, transactivation domain; SET, SET domain. Regions structurally conserved with Trithorax of Drosophila are indicated by the (TRX) label.

minor groove of the DNA, but it recognizes DNA structure domain,38 suggesting that MLL normally functions as a dimer or rather than specific consensus sequences.29 The MT domain is a oligomer. Besides these domains, MLL also has nuclear cysteine-rich CXXC region with homology to a DNA methyl- localization signals in its N-terminal that determine a punctate transferase and methyl-CpG-binding protein 1 (MBD1).29,30 Like localization pattern of the proteins in the nucleus.39 these proteins, the MT domain of MLL has DNA-binding activity to unmethylated CpG sequences, in contrast to MBD1, which binds methylated CpG sequences.31 MLL protein is also known The maturation of MLL protein to have transcriptional repression activity by recruiting repressor complex(es) such as HDAC or polycomb group proteins such as Full-length MLL protein can be cleaved post-translationally by a HPC2 or BMI-1 through MT domain.32 protease, taspase 1, at two conserved amino-acid motifs (CS1: For most proteins with PHD finger domain are components of D/GADD; CS2: D/GVDD).40 Thus, MLL protein is cleaved into chromatin remodeling/transcriptional regulation complexes, at two fragments – an N-termninal 320 kDa (amino acids 1–2666; least some part of chromatin remodeling/transcriptional regula- N320) fragment with transcriptional repression activity and a C- tion activities of normal MLL protein may be associated with terminal 180 kDa (amino acids 2719–3969; C180) fragment PHD fingers.33 In addition, nuclear cyclophilin Cyp33 can bind with strong transcriptional activation properties.41 The N- to the PHD finger of MLL, and overexpression of Cyp33 is terminal fragment comprises a series of AT-hook motifs: the known to have some effects on the expression pattern of HOX SNL1 and SNL2 (speckled nuclear localization signals 1 and 2) genes.34 MLL also has a transcriptional activation domain (TAD) regions; the MT domain, and the PHD zinc-finger domain. The between the PHD fingers and C-terminal SET domain. This C-terminal fragment contains the SET domain and the TAD activation domain of MLL can bind directly to CBP and facilitate domain. The FYRN domain (amino acids 1979–2130) located in the binding of CBP with CREB to promote transcriptional N320 interacts with the FYRC and SET domains (amino acids activation.35 The SET domain is also an evolutionarily highly 3656–3969) located in C180 (Figure 1), thus the two complex conserved region and has activity as a lysine-directed histone MLL peptides build the basic structure required for an MLL methyltransferase that impacts on chromatin structure and multiprotein supercomplex (MLL MPSC)37 that consists of both transcriptional regulation of HOX genes.36,37 Wild-type MLL MLL peptides that confers protein stability and subnuclear contains self-association motifs in the PHD fingers34 and SET localization42 and at least 27 additional proteins.

Leukemia Molecular mechanisms of MLL-associated leukemia Z-Y Li et al 185 The recurrent forms of MLL rearrangements MLL PTDs and MLL gene ampliﬁcations

MLL translocation and MLL fusion partners The so-called PTD leads to fusion of the 50 portion of MLL protein with itself (Figure 1).16 This abnormality is associated In contrast to the majority of other fusion oncoproteins, MLL N with AML, the incidence lying in the range of 3–10% of terminus fuses with a puzzling number of nonrelated proteins, unselected cases45 but rises to 90% of AML cases with trisomy of and they form chimeric proteins. MLL partners can be divided chromosome 11.46 The chimeric transcripts code for proteins as into two groups. The first group is nuclear fusion partners, potent transcriptional activators with potential oncogenic including AF4, AF9, AF10, ENL, ELL, AF17, FKHRL1, AFX1, activity. The finding that partially duplicated forms of MLL also CBP, p300, and so on, which are associated with various aspects had potent coactivator activity suggests that duplication of of transcriptional regulation. The second group is mainly potential DNA binding elements such as the AT hooks or DNA cytoplasmic and frequently associated with cytoskeleton- methyltrasferase homology region may play an important role in depended signal transduction, including AF6, Septin 6, ABI-1, transformation.47 The duplicated DNA binding domains might EEN, FBP17, and so on, possessing structural domains respon- interfere with corepressor binding to CpG repeats, or AT hook- sible for protein–protein interaction (Table 1).13 mediated DNA bending could facilitate transcription factor Despite the heterogeneity of the fusion partners, all known binding further upstream, or increased affinity might stabilize 11q23 translocations delete a large 30 portion of the MLL gene interactions with coactivators.47 A partial nontandem duplica- and connect the remaining 50 part in frame with the tion with the insertion of one AF9 exon that generates three corresponding partner gene. This leads to the expression of a distinct fusion transcripts in B-cell acute lymphoblastic leukemia chimeric protein that has actively transforming properties has been reported.48 (Figure 1). Moreover, as a result of reciprocal translocations, MLL gene amplification was recognized as a recurrent the N-terminal fusion portion of MLL contains two different aberration in AML and MDS, associated with adverse prognosis DNA binding domains (AT-hooks and MT-domain) and the and karyotype complexity.17–19 These patients exhibit intra- critical regions for correct subnuclear localization. Therefore, chromosomal amplification of unrearranged MLL, presumably fusions of the N-terminal portion of the MLL protein with various to facilitate increased MLL expression. Expression analyses partner protein sequences should target the same subnuclear identified MLL as a prominent target of 11q23 amplification and compartment, as evidence from independent studies that have support an etiologic role for MLL gain of function in myeloid demonstrated a nuclear punctate distribution in immunohisto- malignancies.18 chemical studies.39,43 Patients carrying MLL translocations were shown to exhibit a specific gene expression profile of upregulated and downregulated genes, suggesting a novel and The molecular mechanisms of MLL-associated leukemia specific disease phenotype.44 The upregulated genes were some recognized targets of MLL including some HOX genes. There is good evidence that the acquisition of novel properties Deregulated expression of HOX genes typifies certain malig- by the combination of MLL with the fusion partners, rather than nancies. the loss of wild-type MLL function, leads to the generation of an

Table 1 MLL fusion partners

Gene Chromosome locus Functions

Nuclear fusion partner AF4 4q21 Transcriptional activator AF9 9p22 Transcriptional factor AF10 10p12 Transcriptional factor ELL 19p13.1 RNA polymerase II transcription elongation factor ENL 19p13.3 Transcriptional activator AFX Xq13 Forkhead transcriptional factor LAF4 2q11 Transcriptional activator AF3p21 3p21 Not known FKHRL1 6q21 Forkhead transcriptional factor CBP 16p13 Transcriptional coactivator, histone acetylase P300 22q13 Transcriptional coactivator, histone acetylase AF17 17q21 Transcriptional factor, upregulated by b-catenin

Cytoplasmic fusion partner AF6 6q27 Maintenance of cell–cell junctions and cell polarity Septin 6 Xq22 Septin family ABI1 10p11.2 Regulation of endocytosis, cell motility LARG 11q23.3 Activation of Rho GTPases GAS7 17p13 Actin assembly/crosslinking of actin ﬁlaments EEN 19p13.3 Regulation of endocytosis FBP17 9q34 Not known GMPS 3q25 Guanosine monophosphate synthetase GRAF 5q31 Negative regulator of RhoA AF9q34 9q34 Ras GTPase-activating protein GPHN 14q24 Gly and GABA receptors assembly CBL 11q23.3 Negative regulator of receptor tyrosine kinases LASP1 17q21 Not known

Leukemia Molecular mechanisms of MLL-associated leukemia Z-Y Li et al 186 active oncoprotein. Simple truncation of MLL at the site of self-association motifs in some MLL translocation partners has translocation-induced fusion does not lead to development of prompted several investigators to propose that dimerization of leukemias in gene-targeted mice49 or in retroviral transduction/ truncated MLL may be transforming.65–67 There is abundant transplantation models of MLL-associated myeloid leukemogen- indirect evidence for this mechanism. AF10 and AF17 contain esis.50 In a series of knockin studies it was clearly demonstrated leucine zipper motifs that are required for transformation60,67 that MLL needs a fusion partner to become leukemogenic and AF10 is a homotetramer.68 AF3p21 homo-oligomerizes66 whereas a simple truncation did not cause leukemias in and guanine monophosphate synthetase is a tetramer.69 In mice.51,52 Moreover, several structure–function studies testing addition, LCX,70 SEPTING,71 and EEN72 all contain a-helical the transformation assay found critical contribution of the fusion coiled-coil regions that are retained in fusion proteins. These partners.53–55 Recent studies provided more details on MLL coiled-coil self-association domains resemble the rod-like translocations and suggested that different mechanisms might be domains that mediate dimerization of myosin chains and involved in the leukemogenesis by MLL fusion proteins. therefore are also likely to self-associate. Experimental evidence Although the oncogenesis of MLL-associated leukemia is very for dimerization model is the finding that MLL-LacZ knockin complicated, this issue can be dissected from the following mice developed both lymphoid and myeloid leukemias, albeit different aspects. with a long latency period.52 b-Galactosidase has no known transcriptional activity nor is it homologous to any known MLL translocation partner. However, in its active form, the enzyme is Transcriptional activation a tetramer in solution,73 implying that the primary role of the translocation partner in these experimental leukemias is to In many studies, it has been postulated that transcriptional oligomerize MLL. transactivation might be the common denominator of several MLL normally maintains HOX gene transcription in part by fusion partners and transactivational domains are indeed found targeting SET domain methyltransferase activity to HOX gene in some fusion partners. FKHRL1 and AFX are two MLL fusion promoters,36 but this domain and the TAD domain are lost in partners that have well-characterized roles in transcriptional leukemogenic MLL fusion proteins. Forced dimerization/oligo- regulation.56,57 They are members of the forkhead subfamily of merization may alter the association of a DNA binding protein transcription factors (FKHR family) that contain highly con- for its transcriptional cofactors, or the dimerization motifs served forkhead DNA-binding domains (DBDs).58 Transforma- themselves may constitutively recruit transcriptional effector tion mediated by MLL-forkhead fusion proteins required two molecules. Oligomerized chimeras may also sequester essential conserved transcriptional effector domains (CR2 and CR3), each partners or cofactors to exert dominant-negative effects on target of which alone is not sufficient to activate MLL.59 A synthetic gene expression. fusion of MLL with FKHR, a third mammalian forkhead family Moreover, recent experimental data support this MLL member that contains both effector domains, is also capable of dimerization/oligomerization model. Simple dimerization of transforming hematopoietic progenitors in vitro.59 For ENL, the MLL by a cytoplasmic fusion partner (either GAS7 or AF1p) 90 C-terminal amino acids not only constitute a transcriptional activates its transcriptional and oncogenic properties.74 Simi- activator but they are also the minimal component necessary to larly, a coiled-coil dimerization domain of EEN retained in MLL- convert MLL into a leukemogenic oncoprotein.54 Similarly, EEN fusion protein is critical for leukemogenesis caused by MLL- structure/function analysis showed that a highly conserved 82- EEN.72 In addition, MLL-EEN might act as a potential transcrip- amino-acid portion of AF10, comprising two adjacent alpha- tional factor that transactivates the promoter of HOXA7,a helical domains, exhibited transcriptional activation properties potential target gene of MLL.72 Another article showed that the and was sufficient for immortalizing activity when fused to small oligomerization domain of gephyrin converts MLL to an MLL.60 The crucial domain of another MLL fusion partner, ELL, oncogene.75 Forced dimerization of MLL may lead to the is a protein–protein interaction domain that binds to EAF1 (ELL- inappropriate recruitment of accessory factors and create a associated factor).55 EAF1, in turn, comprises a stretch of amino transcriptional activator complex capable of stimulating HOX acids with homology to the transactivator domain of AF4.61 expression in the absence of histone methylation. This model Finally, the MLL partners CBP and p300 are general cofactors in may also account for the oncogenicity of a class of transforming transcriptional activation.62,63 In another report, Zeisig et al64 ‘self-fusion’ MLL proteins, which arise through an intrachromo- provided evidence that a generic transactivator function could somal tandem duplication of N-terminal MLL sequences, replace the authentic MLL fusion partner ENL. These results effectively producing a tethered homodimer of this region fused corroborate the hypothesis that an intrinsic transactivation to the remainder of the MLL coding sequences.47 potential is the common denominator of many MLL fusion partners. Structure/function studies of the respective fusion proteins revealed that the minimal domains required for Chromatin structure changes oncogenesis display transactivation potentials that vary from strong (AFX, FKHR) to weak (ENL, ELL, AF10, CBP). Thus, As discussed above, patients carrying MLL translocations were acquisition of heterologous transcriptional effector domains by shown to exhibit a specific gene expression profile, suggesting a MLL may represent a common oncogenic pathway for dereg- novel and specific disease phenotype. And why? Chromatin ulating its transcriptional functions by some, if not all, of its structure changes of downstream genes essentially account for nuclear fusion partners. the leukemogenesis by MLL fusion proteins. MLL itself is a human homologue of the epigenetic transcriptional regulator Trithorax (TRX) in Drosophila melanogaster.20 MLL is part of Dimerization or oligomerization nuclear regulatory mechanism that establishes an epigenetic transcriptional memory system.76 This system is based on high However, not all MLL translocation partners appear to be molecular weight protein complexes that exert histone/chroma- transcriptional activators. Some are cytoplasmic proteins that tin-modifying/remodeling and transactivating/repressing activ- are unlikely to have a nuclear function. The presence of ities.77 MLL, as a component of MPSC, has been shown that it

Leukemia Molecular mechanisms of MLL-associated leukemia Z-Y Li et al 187 binds directly to HOXA9 and HOXC8 regulatory regions in proapoptotic genes.86 A putative guanine nucleotide exchange chromatin immunoprecipitation assays and by the demonstra- factor (GEF), termed leukemia-associated RhoGEF (LARG), was tion that the SET domain of MLL is a histone H3 (lysine-4)- recently identified upon fusion to the coding sequence of the specific methyltransferase whose activity is associated with MLL gene in AML.87 Although the function of LARG is still HOX gene activation.36,37 As a result, specific gene expression unknown, it exhibits a number of structural domains suggestive patterns will be established and maintained throughout sub- of a role in signal transduction, including a PDZ domain, an LH/ sequent mitotic cell cycles, which in turn will allow cells to RGS domain, and a Dbl homology/pleckstrin homology cope with their cellular fate, or their specific developmental or domain.88 LARG can activate Rho in vivo.87 Furthermore, differentiation pathways. LARG is an integral component of a novel biochemical route However, when translocation occurs, the normal epigenetic whereby G protein-coupled receptors (GPCRs) and heterotri- transcriptional functions of MLL are disrupted and aberrant meric G proteins of the G alpha(12) family stimulate Rho- epigenetic transcriptional functions are contributed by its fusion dependent signaling pathways.88 Taken together, these findings partners. The minimal essential domains of ENL as well as of suggest a potential link between MLL fusion-mediated leuke- AF9 recruit the novel polycomb protein Pc3.78 In addition, mogenesis and the different signal pathways. novel SWI/SNF chromatin-remodeling complexes contain ENL have been identified. The resultant MLL-ENL fusion protein associates and cooperates with SWI/SNF complexes to activate Summary transcription of the promoter of HOXA7.79 Two more fusion partners, ABI1 and AF10, are also connected to ENL either by a As a summary, these different mechanisms may explain some of direct interaction or via the GAS41 protein, an ENL homologue the heterogeneity in MLL-mediated leukemogenesis. For some that interacts with the human SWI/SNF complex.80,81 Polycomb nuclear fusion partners, transcriptional activation may be proteins and SWI/SNF complex in turn are known to be prevailing because acquisition of heterologous transcriptional members of machines that have been connected mostly, but effector domains from these fusion partners will directly not exclusively, with transcriptional silencing. It might be influence downstream gene expression at transcriptional level. possible that, by unknown mechanisms, the assembly of an But for some cytoplasmic fusion partners that are unlikely to MLL-ENL-chromatin remodeling complex leads to chromatin have a nuclear function, dimerization or oligomerization will structure changes that cause a persistent and strong activation of better explain the leukemogenesis by these MLL fusion proteins target genes. Chromatin structure changes maybe are also the because the coiled-coil self-association domains retained in underlying mechanism in CBP and p300 fusion partners MLL fusion proteins may facilitate the dimerization of MLL achieving their transactivation effect. fusion proteins and alter the normal transcriptional regulation activities. Forced dimerization/oligomerization may alter the association between MLL and its transcriptional cofactors, or the Association with signal transduction dimerization motifs themselves may constitutively recruit transcriptional effector molecules or accessory factors and SH3 (Src homology 3) domain, a well-known domain contained create a transcriptional activator complex exert dominant- in many proteins intermediating signal transduction, plays an negative effects on target gene expression. important and intriguing role in MLL fusion partners. EBP, a Furthermore, MLL fusion partners will result in the remodeling novel EEN binding protein, interacts with the SH3 domain of of MPSC, the altered MPSC will recruit some accessory factors EEN through a proline-rich motif PPERP.82 EBP is a ubiquitous that may influence gene expression at transcriptional level or protein that is normally expressed in the cytoplasm but is influence signal transduction in expanded range. Due to the recruited to the nucleus by MLL-EEN with a punctate localiza- heterogeneity of MLL fusion partners, the components of MPSC tion pattern characteristic of the MLL chimeric proteins.82 EBP alter from one to another and different mechanisms will mediate interacts simultaneously with EEN and Sos, a guanine-nucleo- the oncogenesis of MLL-associated leukemias. tide exchange factor for Ras. Coexpression of EBP with EEN Though mechanisms of leukemogenesis by MLL fusion leads to suppression of Ras-induced cellular transformation and proteins are various, there is the same result – the alteration of Ras-mediated activation of Elk-1.82 Taken together, these gene expression profiles. Certainly, different MLL fusion proteins findings suggest a new mechanism for MLL-EEN-mediated will result in different gene expression profiles, but the leukemogenesis in which MLL-EEN interferes with the Ras- establishment of these expression profiles is ultimately achieved suppressing activities of EBP through direct interaction. More- through the epigenetic regulation. over, SH3 domains of EEN and ABI-1 can interact with different proline-rich domains of synaptojanin,83 a member of the inositol phosphatase family that has recently been shown to regulate cell Conclusions and perspective proliferation and survival. Similarly, the AF3p21 gene encodes a protein consisting of 722 amino acids, which has an SH3 Different mechanisms may be involved in the leukemogenesis domain, a proline-rich domain, and a bipartite nuclear by different aberrant MLL proteins and there is some connection localization signal. The structural characteristics suggest the between these mechanisms: (1) As discussed above, dimeriza- possibility that AF3p21 protein plays a role in signal transduction (or oligomerization) might be the common mechanism for tion in the nucleus.84 Another MLL fusion partner AF17 was leukemogenesis by aberrant MLL proteins derived from different identified as a downstream gene of the b-catenin/T-cell factor MLL rearrangements (including MLL fusion with some cytoplas- pathway. AF17 gene product is likely to be involved in the mic partners, MLL PTD or MLL gene amplification);47(2) b-catenin-T-cell factor/lymphoid enhancer factor signaling aberrant MLL MPSC will recruit some chromatin-remodeling pathway and to function as a growth-promoting, oncogenic complexes that result in chromatin structure changes and even protein.85 Similarly, FKHRL1 and AFX are downstream targets aberrant gene expression profiles; (3) some MLL fusion proteins of protein kinase B (PKB/Akt) in a conserved signaling derived from MLL fusion with some cytoplasmic partners can act pathway that regulates expression of cell cycle regulatory and as transcription regulator via dimerization (eg MLL-EEN);72

Leukemia Molecular mechanisms of MLL-associated leukemia Z-Y Li et al 188

Figure 2 Multiple roles of normal and aberrant MLL proteins in development programming. Normal MLL protein is required for maintaining for normal gene expression profile and normal development programming. However, aberrant MLL proteins derived from MLL translocation, MLL partial tandem duplication or MLL gene amplification will result in aberrant gene expression profile and aberrant development programming and eventually will result in leukemia. Different mechanisms are involved in leukemogenesis by aberrant MLL proteins and there are some connections among them.

(4) some fusion partners may link some signal molecules to MLL by a grant from the National Natural Science Foundation of China MPSC and may result in extensive alteration in gene expression (N0. 30393110). profiles. Thus, the aberrant MLL proteins presumably affect normal cellular control mechanisms, including signal transduction, cell cycle, and so on, possibly by a direct action on its References target genes (eg HOX gene) expression occurring at the level of chromatin-mediated activation or by an indirect action on other 1 Cox MC, Panetta P, Lo-Coco F, Del Poeta G, Venditti A, Maurillo L nontargeted genes (Figure 2). et al. Chromosomal aberration of the 11q23 locus in acute leukemia and frequency of MLL gene translocation: results in 378 In order to study leukemogenesis of aberrant MLL proteins, adult patients. Am J Clin Pathol 2004; 122: 298–306. loss-of-function models and gain-of-function models are often 2 del Mar Bellido M, Nomdedeu JF. Adult de novo acute myeloid put to use to analyze gene expression patterns. Dysregulation of leukemias with MLL rearrangements. Leuk Res 1999; 23: 585–588. HOX gene family members is implicated as a dominant 3 Stock W, Thirman MJ, Dodge RK, Rowley JD, Diaz MO, Wurster- mechanism of leukemic transformation induced by chimeric Hill D et al. Detection of MLL gene rearrangements in adult acute MLL oncogenes.89 The strategy of comparing patterns of gene lymphoblastic leukemia. A Cancer and Leukemia Group B Study. Leukemia 1994; 8: 1918–1922. expression in loss-of-function models to gain-of-function models 4 Sorensen PH, Chen CS, Smith FO, Arthur DC, Domer PH, and leukemic cells harboring MLL fusion (or MLL partial Bernstein ID et al. Molecular rearrangements of the MLL gene duplication or MLL amplification) oncogenes will not only help are present in most cases of infant acute myeloid leukemia and are us understand globally the mechanism by which aberrant MLL strongly correlated with monocytic or myelomonocytic pheno- oncogenes affect the normal cellular processes but also provide types. J Clin Invest 1994; 93: 429–437. more details in antileukemia therapies. To clarify the hetero- 5 Martinez-Climent JA, Thirman MJ, Espinosa III R, Le Beau MM, Rowley JD. Detection of 11q23/MLL rearrangements in infant geneity of leukemogenesis by different aberrant MLL proteins leukemias with fluorescence in situ hybridization and molecular derived from different chromosomal rearrangements, investiga- analysis. Leukemia 1995; 9: 1299–1304. tion on diversification of MPSC components will provide more 6 Satake N, Maseki N, Nishiyama M, Kobayashi H, Sakurai M, Inaba information to us. As discussed above, some evidence suggests a H et al. Chromosome abnormalities and MLL rearrangements potential link between MLL fusion-mediated leukemogenesis in acute myeloid leukemia of infants. Leukemia 1999; 13: and the different signal pathways. To testify this possibility, 1013–1017. 7 Andersen MK, Christiansen DH, Jensen BA, Ernst P, Hauge G, study on MPSC composition combined with the information Pedersen-Bjergaard J. Therapy-related acute lymphoblastic leu- from the study of signal transduction will be helpful. kaemia with MLL rearrangements following DNA topoisomerase II inhibitors, an increasing problem: report on two new cases and review of the literature since 1992. Br J Haematol 2001; Acknowledgements 114: 539–543. 8 Super HJ, McCabe NR, Thirman MJ, Larson RA, Le Beau MM, Pedersen-Bjergaard J et al. Rearrangements of the MLL gene in We thank Dr Gong-Hong Wei and other colleagues for helpful therapy-related acute myeloid leukemia in patients previously discussion and critical reading of the manuscript. We apologize treated with agents targeting DNA-topoisomerase II. Blood 1993; for any errors or omissions in this review. This work was supported 82: 3705–3711.

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