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Recurrent translocation (10;17)(p15; q21) in acute poorly diff erentiated myeloid leukemia likely results in ZMYND11-MBTD1 fusion

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Leukemia & Lymphoma
Authors:
Etienne De Braekeleer at AstraZeneca
Etienne De Braekeleer
Régis Auffret
Régis Auffret
Régis Auffret
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Nathalie Douet-Guilbert at Centre Hospitalier Universitaire de Brest
Nathalie Douet-Guilbert
Audrey Basinko
Audrey Basinko
Audrey Basinko
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
Leukemia & Lymphoma, May 2014; 55(5): 1189–1190
© 2014 Informa UK, Ltd.
ISSN: 1042-8194 print / 1029-2403 online
DOI: 10.3109/10428194.2013.820292
*
Present address: Division of Stem Cells and Cancer, Heidelberg Institute for Stem Cell Technology and Experimental Medicine (HI-STEM), German Cancer
Research Center (DKFZ), Heidelberg, Germany
Correspondence: Pr. Marc De Braekeleer, Laboratoire de Cytog é n é tique, H ô pital Morvan, b â timent 5bis, CHRU Brest, 2, avenue Foch, F-29609 Brest cedex,
France. Tel: ⫹ 33(0)2-98-22-36-94. Fax: ⫹ 33(0)2-98-22-39-61. E-mail: marc.debraekeleer@univ-brest.fr
Received 11 April 2013 ; revised 14 May 2013 ; accepted 23 June 2013
LETTER TO THE EDITOR
Recurrent translocation (10;17)(p15;q21) in acute poorly diff erentiated
myeloid leukemia likely results in ZMYND11 – MBTD1 fusion
Etienne De Braekeleer
1
*
, R é g i s A u ff ret
1
, Nathalie Douet-Guilbert
1,2,3
, Audrey Basinko
3
,
Marie-Jos é e Le Bris
3
, Fr é d é ric Morel
1,2,3
& Marc De Braekeleer
1,2,3
1
Laboratoire d ’ Histologie, Embryologie et Cytog é n é tique, Facult é de M é decine et des Sciences de la Sant é , Universit é de
Bretagne Occidentale, Brest, France,
2
Institut National de la Sant é et de la Recherche M é dicale (INSERM) U1078, Brest, France
and
3
Service de Cytog é n é tique, Cytologie et Biologie de la Reproduction, H ô pital Morvan, CHRU Brest, Brest, France
Translocation (10;17)(p15;q21) is a recurrent abnormality
that has been reported in only seven cases of acute leukemia,
including two by our group [1] (available at: http://cgap.nci.
nih.gov/Chromosomes/Mitelman, accessed April 2013).  is
translocation appears to be highly speci cally associated
with poorly di erentiated acute myeloid leukemia. Indeed,
two patients had acute myeloid leukemia with minimal dif-
ferentiation (French – American – British [FAB] type M0) [2,3]
and four acute myeloid leukemia without maturation (FAB
type M1) [1,4,5]. A sole patient had acute pre-B lymphoblas-
tic leukemia (FAB type L1) [6].
Although band 17q21 is rich in candidate genes involved
in leukemogenesis, only the retinoic acid receptor alpha
( RARA ) gene is known to be rearranged in acute promyelo-
cytic leukemia. Genes involved in other types of leukemia
remain to be found. We showed previously that RARA was
not rearranged in two patients with t(10;17)(p15;q21) but
remained on the derivative chromosome 17, suggesting that
the breakpoint involved in this translocation was telomeric
to its locus [1]. Here, we present the results of  uorescence in
situ hybridization (FISH) using BAC (bacterial arti cial chro-
mosome) clones to identify the candidate genes involved in
both patients.
Patient 1, a 13-year-old boy, was  rst seen because of
persistent fever and asthenia. A diagnosis of acute myeloid
leukemia, M1 subtype in the FAB classi cation, was made.
Induction therapy followed by three consolidation courses
led to a complete remission (CR); the patient is still in CR 71
months following diagnosis. Patient 2, a 40-year-old woman,
was  rst seen because of a history of asthenia and dorsal
and leg pain. A diagnosis of AML, M1 subtype, was made.
e patient was treated with induction therapy followed by
consolidation therapy.  e patient achieved two CRs and
received two bone marrow transplants. She died 37 months
following the initial diagnosis. Clinical and laboratory data
on both patients were previously reported in detail [1].
Cytogenetic analysis was performed on bone marrow
cells cultured for 24 h and synchronized with  uorodeoxy-
uridine (FUdR) at the time of diagnosis and during evolution.
e chromosomes were RHG-banded and the karyotype
described according to the International System for Human
Cytogenetic Nomenclature (ISCN 2009) [7]. In patient 1,
14 of the 23 metaphases observed at diagnosis showed an
abnormal karyotype: 46,XY,t(10;17)(p15;q21)[5]/47,XY,ide
m, ⫹ 13[9]/46,XY[9]. In patient 2, at diagnosis, nine of the 25
metaphases showed an abnormal karyotype: 46,XX,t(10;17)
(p15;q21)[3]/46,XX,idem,der(11)(11pter → 11q13::11q23 → 11
q14::13q34 → 13qter), der(13)(13pter → 13q34::11q23 → 11qter)
[6]/46,XX[16].  e sole translocation (10;17) was found dur-
ing both relapses.
FISH analyses using BAC libraries were then used to
clone the translocation breakpoints on chromosomes 10
and 17, as previously described [8]. We identi ed the BAC
clones of interest through the human genome browser
database of the genome bioinformatics group at the Uni-
versity of California at Santa Cruz (UCSC; http://genome.
ucsc.edu/).  ey were then ordered on the site of the Chil-
dren ’ s Hospital Oakland Research Institute in Oakland,
CA (http://bacpac.chori.org/). In a  rst step, we used BAC
clones covering bands 10p15 and 17q21, spaced every 1.5 – 2
Mb. Once the boundaries of both breakpoint regions were
determined, overlapping BAC clones were ordered to re ne
the regions.
BAC clones RP11-10D13, located in band 10p15.3, and
RP11-379D19, located in band 17q21.33, were shown to be
split between der(10) and der(17). Both breakpoints were
further re ned with overlapping BAC clones.  is allowed
us to assign the breakpoint on chromosome 10 between
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 E. De Braekeleer et al.
positions 288 915 and 322 071 [based on the UCSC Genome
Browser on Human February 2009 (GRCh37/hg19) Assem-
bly]; this region contains the telomeric part of the ZMYND11
(zinc  nger, MYND-type containing 11) gene.  e breakpoint
on chromosome 17 was located between positions 49 322
093 and 49 322 113; this region contains the telomeric part of
MBDT1 (mbt domain containing 1) gene. Co-hybridization
with RP11-387K19 (containing the ZMYND11 gene) and
RP11-326B24 (containing the MBDT1 gene) clones showed
co-localization of both probes in abnormal leukemia cells
from both patients.
e ZMYND11 (alias BS69 ) gene contains 15 exons, of
which 14 are coding, spanning 120 kb. Di erent isoforms
are generated by alternatively spliced transcript variants.
ZMYND11 localizes to the nucleus and contains three motifs
involved in transcription regulation: a PHD  nger and bro-
modomain in its N-terminal half, and a MYND domain at its
C terminus [9,10].
e MYND (Myeloid, Nervy and DEAF-1) domain is simi-
lar to that found in the ETO/MTG8 protein, which is fused to
RUNX1 in acute myelogenous leukemia [11]. It is a conserved
two-zinc  nger motif present in a large group of proteins. Full
transcriptional repression by ZMYND11 requires the MYND
domain, which interacts with the N-CoR/mSin3/HDAC1
complex that causes transcriptional repression [10,12,13].
e MYND domain was also shown to interact with other
proteins, including the PxLxP motif in E1A, the Epstein – Barr
virus oncoprotein EBNA2 and MGA (a MYC-related cellu-
lar transcription factor) [14]. Furthermore, ZMYND11 also
inhibits the transcriptional activity of MYB [15].  erefore,
ZMYND11 could have tumor suppressor-like properties, by
down-regulating transcription factors that have oncogenic
potential.
e MBTD1 gene contains 17 exons, of which 15 are cod-
ing, spanning 82 kb. MBTD1 localizes to the nucleus and con-
tains a FCS-type zinc  nger at the N-terminus with putative
regulatory function, and four MBT (malignant brain tumor)
repeats at the C-terminus [16]. MBTD1 is a putative Polycomb
group protein, sharing homologies with L3MBTL1, L3MBTL2
and L3MBTL3 [17 – 19]. Proteins belonging to the Polycomb
group maintain the transcriptionally repressive state of genes,
probably via chromatin remodeling [20]. MBTD1, L3MBTL1
and L3MBTL3 were implicated in hematopoiesis [21,22].
Unfortunately, no RNA was available to determine
whether a fusion transcript was generated as a consequence
of the translocation. However, it is possible that loss of the
MYND domain induced a lack of transcriptional repression
and tumor suppressor-like properties of the ZMYND11 pro-
tein. Four other cases of t(10;17)(p15;q21)-associated AML
without or with minimal maturation were reported in the lit-
erature. It remains to be determined whether the same genes
are involved in these patients. More studies are necessary to
determine whether a ZMYND11 – MBTD1 fusion transcript is
generated and to analyze the functional consequences of the
t(10;17)(p15;q21) translocation.
Potential confl ict of interest: Disclosure forms provided
by the authors are available with the full text of this article at
www.informahealthcare.com/lal.
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For personal use only.
... A specific recurrent translocation, t(10;17) (p15;q21), has been described over the past few years and has been linked to a cannibalistic form of AML (Plesa and Sujobert, 2019). Based on FISH analyses, this translocation creates a merge of ZMYND11 and MBTD1 genes (De Braekeleer et al., 2013). The break points are in exon 11 or 12 of ZMYND11 and exon 3 of MBTD1, creating an in-frame coding sequence that encompasses a truncated ZMYND11 fused to a full-length MBTD1 (De Braekeleer et al., 2013;de Rooij et al., 2016;Plesa and Sujobert, 2019;Tempescul et al., 2007;Yamamoto et al., 2018) (Fig. 1A). ...
... Based on FISH analyses, this translocation creates a merge of ZMYND11 and MBTD1 genes (De Braekeleer et al., 2013). The break points are in exon 11 or 12 of ZMYND11 and exon 3 of MBTD1, creating an in-frame coding sequence that encompasses a truncated ZMYND11 fused to a full-length MBTD1 (De Braekeleer et al., 2013;de Rooij et al., 2016;Plesa and Sujobert, 2019;Tempescul et al., 2007;Yamamoto et al., 2018) (Fig. 1A). ...
Oncogenic ZMYND11-MBTD1 fusion protein anchors the NuA4/TIP60 histone acetyltransferase complex to the coding region of active gene
Preprint
Full-text available
  • Mar 2021
A chromosomal translocation found in cannibalistic acute myeloid leukemia (AML) leads to an in-frame fusion of the transcription elongation repressor ZMYND11 to MBTD1, a subunit of the NuA4/TIP60 histone acetyltransferase (HAT) complex. In contrast to the NuA4/TIP60 complex, ZMYND11 is linked to repression of actively transcribed genes through recognition of H3.3K36me3. To understand the abnormal molecular events that expression of this ZMYND11-MBTD1 fusion protein can create, we performed its biochemical and functional characterization in comparison to each individual fusion partner. ZMYND11-MBTD1 is stably incorporated into the endogenous NuA4/TIP60 complex but does not bring any additional interactors as the fusion lacks the MYND domain of ZMYND11. Nevertheless, this truncated ZMYND11 moiety in the fusion mostly leads to mislocalization of the NuA4/TIP60 complex on the body of genes normally bound by ZMYND11 in the genome. This can be correlated to increased chromatin acetylation and altered gene transcription, most notably on the MYC oncogene. Importantly, expression of ZMYND11-MBTD1, but not the individual fusion partners, during embryonic stem cell differentiation, leads to decreased expression of specific differentiation markers, while favoring Myc-driven pluripotency. Altogether, these results indicate that the ZMYND11-MBTD1 fusion protein functions primarily by mistargeting the NuA4/TIP60 complex to the body of genes, altering normal transcription of specific genes, likely driving oncogenesis in part through the Myc regulatory network. Highlights - A recurrent chromosomal translocation detected in cannibalistic acute myeloid leukemia leads to the production of a ZMYND11-MBTD1 fusion protein. - The ZMYND11-MBTD1 fusion protein is stably incorporated into the endogenous NuA4/TIP60 complex - ZMYND11-MBTD1 leads to mistargeting of NuA4-TIP60 activity to the coding region of ZMYND11-target genes, altering gene expression and transcript isoforms. - ZMYND11-MBTD1 binds the MYC gene leading to its upregulation, favoring growth and pluripotency while inhibiting differentiation markers.
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... P ost-translational modification (PTM) of histones provides a fundamental means for modulating gene expression and determining cellular identities during development and cell differentiation, and its deregulation is intimately associated with pathogenesis of human cancers, including acute myeloid leukemia (AML) [1][2][3][4][5][6] . Recently, a new recurrent chromosomal translocation t(10;17)(p15;q21) was detected among a subset of AML patients, which produces an abnormal chimeric gene by fusing an N-terminal gene segment (i.e., exons 1-11 or 1-12) of Zinc Finger MYND-Type Containing 11 (ZMYND11) in-frame with the entire coding region (exons [3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] of Malignant Brain Tumor domain containing 1 (MBTD1) [7][8][9][10][11] . The resultant chimeric protein termed ZMYND11-MBTD1 (hereafter referred to as "ZM") harbors the PHD, Bromo and PWWP domains of ZMYND11 and full-length MBTD1 (Fig. 1a). ...
... P ost-translational modification (PTM) of histones provides a fundamental means for modulating gene expression and determining cellular identities during development and cell differentiation, and its deregulation is intimately associated with pathogenesis of human cancers, including acute myeloid leukemia (AML) [1][2][3][4][5][6] . Recently, a new recurrent chromosomal translocation t(10;17)(p15;q21) was detected among a subset of AML patients, which produces an abnormal chimeric gene by fusing an N-terminal gene segment (i.e., exons 1-11 or 1-12) of Zinc Finger MYND-Type Containing 11 (ZMYND11) in-frame with the entire coding region (exons [3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] of Malignant Brain Tumor domain containing 1 (MBTD1) [7][8][9][10][11] . The resultant chimeric protein termed ZMYND11-MBTD1 (hereafter referred to as "ZM") harbors the PHD, Bromo and PWWP domains of ZMYND11 and full-length MBTD1 (Fig. 1a). ...
ZMYND11-MBTD1 induces leukemogenesis through hijacking NuA4/TIP60 acetyltransferase complex and a PWWP-mediated chromatin association mechanism
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Recurring chromosomal translocation t(10;17)(p15;q21) present in a subset of human acute myeloid leukemia (AML) patients creates an aberrant fusion gene termed ZMYND11-MBTD1 (ZM); however, its function remains undetermined. Here, we show that ZM confers primary murine hematopoietic stem/progenitor cells indefinite self-renewal capability ex vivo and causes AML in vivo. Genomics profilings reveal that ZM directly binds to and maintains high expression of pro-leukemic genes including Hoxa, Meis1, Myb, Myc and Sox4. Mechanistically, ZM recruits the NuA4/Tip60 histone acetyltransferase complex to cis-regulatory elements, sustaining an active chromatin state enriched in histone acetylation and devoid of repressive histone marks. Systematic mutagenesis of ZM demonstrates essential requirements of Tip60 interaction and an H3K36me3-binding PWWP (Pro-Trp-Trp-Pro) domain for oncogenesis. Inhibitor of histone acetylation-‘reading’ bromodomain proteins, which act downstream of ZM, is efficacious in treating ZM-induced AML. Collectively, this study demonstrates AML-causing effects of ZM, examines its gene-regulatory roles, and reports an attractive mechanism-guided therapeutic strategy.
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... Here we demonstrated a potential pathogenic role in SCZ for a previously undescribed DNM in ZMYND11. Importantly, this gene has been implicated in tumorigenesis (De Braekeleer et al., 2014;Li et al., 2021) and syndromic intellectual disability, based on its biological functions and the previously identified mutations. Its role in tumor formation has been studied extensively and is connected to the loss of its co-repressor function of actively transcribed genes (Chen et al., 2019). ...
View
... MBTD1 mutations have been found in endometrial stromal sarcoma and leukemia [26,27]. In prostate cancer, MBTD1 was suggested to be linked to increased prostatespecific antigen level, tumor metastasis, aggressive clinicopathological characteristics, and poor survival of patients [15]. ...
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... However, ZMYND11 is a significant contributor to cancer development. For example, copy number variations of ZMYND11 were found in clinical samples of patients with several types of hematological malignancies [268], and it is fused to the malignant brain tumor domain containing 1 (MBTD1) protein via a chromosomal translocation associated with acute myeloid leukemia [269,270]. ZMYND11 has been shown to specifically bind H3.3K36me3, and its histone-binding activity is sensitive to changes in amino acid substitutions in the histone tail as well as adjacent PTMs [271]. Histone H3.3 contains a serine at position 31, and the replacement of this residue with an alanine in histone H3.1 and H3.2, or phosphorylation of serine 31 in histone H3.3, significantly weakened the bind binding interaction [271]. ...
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  • May 2021
  • NAT REV CANCER
The genetic information of human cells is stored in the context of chromatin, which is subjected to DNA methylation and various histone modifications. Such a ‘language’ of chromatin modification constitutes a fundamental means of gene and (epi)genome regulation, underlying a myriad of cellular and developmental processes. In recent years, mounting evidence has demonstrated that miswriting, misreading or mis-erasing of the modification language embedded in chromatin represents a common, sometimes early and pivotal, event across a wide range of human cancers, contributing to oncogenesis through the induction of epigenetic, transcriptomic and phenotypic alterations. It is increasingly clear that cancer-related metabolic perturbations and oncohistone mutations also directly impact chromatin modification, thereby promoting cancerous transformation. Phase separation-based deregulation of chromatin modulators and chromatin structure is also emerging to be an important underpinning of tumorigenesis. Understanding the various molecular pathways that underscore a misregulated chromatin language in cancer, together with discovery and development of more effective drugs to target these chromatin-related vulnerabilities, will enhance treatment of human malignancies. Deregulation of chromatin modification underlies a myriad of oncogenic processes. This Review synthesizes the many connections between chromatin modifications and cancer, discussing recent advances and highlighting options for therapeutic targeting.
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Using Bacterial Artificial Chromosomes in Leukemia Research: The Experience at the University Cytogenetics Laboratory in Brest, France
Article
Full-text available
  • Jan 2011
  • J BIOMED BIOTECHNOL
The development of the bacterial artificial chromosome (BAC) system was driven in part by the human genome project in order to construct genomic DNA libraries and physical maps for genomic sequencing. The availability of BAC clones has become a valuable tool for identifying cancer genes. We report here our experience in identifying genes located at breakpoints of chromosomal rearrangements and in defining the size and boundaries of deletions in hematological diseases. The methodology used in our laboratory consists of a three-step approach using conventional cytogenetics followed by FISH with commercial probes, then BAC clones. One limitation to the BAC system is that it can only accommodate inserts of up to 300 kb. As a consequence, analyzing the extent of deletions requires a large amount of material. Array comparative genomic hybridization (array-CGH) using a BAC/PAC system can be an alternative. However, this technique has limitations also, and it cannot be used to identify candidate genes at breakpoints of chromosomal rearrangements such as translocations, insertions, and inversions.
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L3MBTL1 polycomb protein, a candidate tumor suppressor in del(20q12) myeloid disorders, is essential for genome stability
Article
Full-text available
  • Dec 2010
  • P NATL ACAD SCI USA
The l3mbtl1 gene is located on the long arm of chromosome 20 (q12), within a region commonly deleted in several myeloid malignancies. L3MBTL1 is a human homolog of the Drosophila polycomb L(3)MBT tumor suppressor protein and thus a candidate tumor suppressor in del(20q12) myeloid disorders. We used the loss-of-function approach to explore the possible tumor suppressive mechanism of L3MBTL1 and found that depletion of L3MBTL1 from human cells causes replicative stress, DNA breaks, activation of the DNA damage response, and genomic instability. L3MBTL1 interacts with Cdc45, MCM2-7 and PCNA, components of the DNA replication machinery, and is required for normal replication fork progression, suggesting that L3MBTL1 causes DNA damage, at least in part, by perturbing DNA replication. An activated DNA damage response and genomic instability are common features in tumorigenesis and a consequence of overexpression of many oncogenes. We propose that the loss of L3MBTL1 contributes to the development of 20q(-) hematopoietic malignancies by inducing replicative stress, DNA damage, and genomic instability.
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Structural studies of a four-MBT repeat protein MBTD1
Article
Full-text available
The Polycomb group (PcG) of proteins is a family of important developmental regulators. The respective members function as large protein complexes involved in establishment and maintenance of transcriptional repression of developmental control genes. MBTD1, Malignant Brain Tumor domain-containing protein 1, is one such PcG protein. MBTD1 contains four MBT repeats. We have determined the crystal structure of MBTD1 (residues 130-566aa covering the 4 MBT repeats) at 2.5 A resolution by X-ray crystallography. The crystal structure of MBTD1 reveals its similarity to another four-MBT-repeat protein L3MBTL2, which binds lower methylated lysine histones. Fluorescence polarization experiments confirmed that MBTD1 preferentially binds mono- and di-methyllysine histone peptides, like L3MBTL1 and L3MBTL2. All known MBT-peptide complex structures characterized to date do not exhibit strong histone peptide sequence selectivity, and use a "cavity insertion recognition mode" to recognize the methylated lysine with the deeply buried methyl-lysine forming extensive interactions with the protein while the peptide residues flanking methyl-lysine forming very few contacts [1]. Nevertheless, our mutagenesis data based on L3MBTL1 suggested that the histone peptides could not bind to MBT repeats in any orientation. The four MBT repeats in MBTD1 exhibits an asymmetric rhomboid architecture. Like other MBT repeat proteins characterized so far, MBTD1 binds mono- or dimethylated lysine histones through one of its four MBT repeats utilizing a semi-aromatic cage. This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.
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Methylation-state-specific recognition of histones by the MBT repeat protein L3MBTL2
Article
Full-text available
  • Mar 2009
  • NUCLEIC ACIDS RES
The MBT repeat has been recently identified as a key domain capable of methyl-lysine histone recognition. Functional work has pointed to a role for MBT domain-containing proteins in transcriptional repression of developmental control genes such as Hox genes. In this study, L3MBTL2, a human homolog of Drosophila Sfmbt critical for Hox gene silencing, is demonstrated to preferentially recognize lower methylation states of several histone-derived peptides through its fourth MBT repeat. High-resolution crystallographic analysis of the four MBT repeats of this protein reveals its unique asymmetric rhomboid architecture, as well as binding mechanism, which preclude the interaction of the first three MBT repeats with methylated peptides. Structural elucidation of an L3MBTL2-H4K20me1 complex and comparison with other MBT-histone peptide complexes also suggests that an absence of distinct surface contours surrounding the methyl-lysine-binding pocket may underlie the lack of sequence specificity observed for members of this protein family.
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BS69, a novel adenovirus E1A-associated protein that inhibits E1A transactivation
Article
Full-text available
  • Aug 1995
The adenovirus E1A gene products are nuclear phosphoproteins that can transactivate the other adenovirus early genes as well as several cellular genes, and can transform primary rodent cells in culture. Transformation and transactivation by E1A proteins is most likely to be mediated through binding to several cellular proteins, including the retinoblastoma gene product pRb, the pRb-related p107 and p130, and the TATA box binding protein TBP. We report here the cloning of BS69, a novel protein that specifically interacts with adenovirus 5 E1A. BS69 has no significant homology to known proteins and requires the region that is unique to the large (289R) E1A protein for high affinity binding. BS69 and E1A proteins coimmunoprecipitate in adenovirus-transformed 293 cells, indicating that these proteins also interact in vivo. BS69 specifically inhibits transactivation by the 289R E1A protein, but not by the 243R E1A protein. BS69 also suppressed the E1A-stimulated transcription of the retinoic acid receptor in COS cells, but did not affect the cellular E1A-like activity that is present in embryonic carcinoma cells. Our data indicate that BS69 is a novel and specific suppressor of E1A-activated transcription.
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The MYND Motif Is Required for Repression of Basal Transcription from the Multidrug Resistance 1 Promoter by the t(8;21) Fusion Protein
Article
Full-text available
  • Jul 1998
Chromosomal translocations in acute leukemia that affect the AML-1/CBFβ transcription factor complex create dominant inhibitory proteins. However, the mechanisms by which these proteins act remain obscure. Here we demonstrate that the multidrug resistance 1 (MDR-1) promoter is a target for AML/ETO transcriptional repression. This repression is of basal, not activated, expression from the MDR-1 promoter and thus represents a new mechanism for AML/ETO function. We have defined two domains in AML/ETO that are required for repression of basal transcription from the MDR-1 promoter: a hydrophobic heptad repeat (HHR) motif and a conserved zinc finger (ZnF) domain termed the MYND domain. The HHR mediates formation of AML/ETO homodimers and AML/ETO-ETO heterodimers. Single serine substitutions at conserved cysteine residues within the predicted ZnFs also abrogate transcriptional repression. Finally, we observe that AML/ETO can also inhibit Ets-1 activation of the MDR-1 promoter, indicating that AML/ETO can disrupt both basal and Ets-1-dependent transcription. The fortuitous inhibition of MDR-1 expression in t(8;21)-containing leukemias may contribute to the favorable response of these patients to chemotherapeutic drugs.
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Histone Recognition by Human Malignant Brain Tumor Domains
Article
  • Sep 2012
  • J MOL BIOL
Histone methylation has emerged as an important covalent modification involved in a variety of biological processes, especially regulation of transcription and chromatin dynamics. Lysine methylation is found in three distinct states (monomethylation, dimethylation and trimethylation), which are recognized by specific protein domains. The malignant brain tumor (MBT) domain is one such module found in several chromatin regulatory complexes including Polycomb repressive complex 1. Here, we present a comprehensive characterization of the human MBT family with emphasis on histone binding specificity. SPOT-blot peptide arrays were used to screen for the methyllysine-containing histone peptides that bind to MBT domains found in nine human proteins. Selected interactions were quantified using fluorescence polarization assays. We show that all MBT proteins recognize only monomethyllysine and/or dimethyllysine marks and provide evidence that some MBT domains recognize a defined consensus sequence while others bind in a promiscuous, non-sequence-specific manner. Furthermore, using structure-based mutants, we identify a triad of residues in the methyllysine binding pocket that imparts discrimination between monomethyllysine and dimethyllysine. This study represents a comprehensive analysis of MBT substrate specificity, establishing a foundation for the rational design of selective MBT domain inhibitors that may enable elucidation of their role in human biology and disease.
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Abnormalities of the short arm of chromosome 9 with partial loss of material in hematologic disorders
Article
  • Aug 1987
Clinical, hematological, and cytogenetic data of 32 patients with loss of part of the short arm of chromosome 9 (9p-) are reviewed. There were 20 acute lymphoblastic leukemia (ALL), seven non-Hodgkin lymphoma (NHL), three acute myeloid leukemia, one refractory anemia with excess blasts in transformation, and one chronic myeloid leukemia (CML) in blast crisis. The cytogenetic findings were heterogeneous: 13 cases of del(9)(p21), among them four as sole karyotypic change; five cases of del(9)(p12), three of them as sole karyotypic change; four patients with i(9q), three with unbalanced translocations involving 9p12; and seven with unbalanced translocations involving 9p21. In addition, 10 patients showed known specific translocations for determined subgroups of ALL, NHL, and CML. The immunological phenotypes in the 20 ALL patients were common ALL (35%), pre-B-ALL (35%), B-ALL (5%), T-ALL (15%), and null ALL (10%). Three NHL were of T cell origin and the others of B cell origin. No specific association between the karyotypic change, immunophenotype, and clinical presentation could be ascertained for patients with ALL, acute myeloid leukemia, CML in blast crisis, and B-NHL. In T-NHL, three children with deletion of 9p, T immunoblastic lymphoma originating from common thymocyte and presenting with a mediastinal mass and pleural effusion may constitute a definite subgroup with good prognosis. All other cases had a poor outcome. Previously suggested association of 9p- with T-ALL and "lymphomatous features" was not confirmed.
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Structure and expression of the human MTG8/ETO gene
Article
  • Jun 1998
  • GENE
Translocations involving the putative proto-oncogene MTG8/ETO on 8q22 are frequently found in acute myeloid leukemia. To date, little is known of the genomic organization of this gene. Here, we report that the MTG8 gene consists of 13 exons distributed over 87 kb of genomic DNA. Two polymorphic microsatellite repeats are described, including one in intron 3 (three alleles; heterozygosity 0.34) and another in the 3'UTR (15 alleles; heterozygosity 0.89). Expression of MTG8 was detected in a variety of normal human tissues with the highest mRNA levels occurring in brain and heart. Previously, two mRNA forms produced by the alternative usage of the first exon have been reported. We now describe a novel, abundantly expressed, alternatively spliced transcript resulting from the inclusion of a 155-bp exon (designated 9a) that changes the reading frame and introduces a premature stop codon. Identical alternatively spliced mRNA variants were found to be produced by the highly conserved homologous gene (Cbfa2t1) in the mouse, suggesting an evolutionary significance.
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