Kojo S J Elenitoba-Johnson

Concordia University–Ann Arbor, Ann Arbor, Michigan, United States

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Publications (113)620.18 Total impact

  • [show abstract] [hide abstract]
    ABSTRACT: Deregulation of signaling pathways controlled by protein phosphorylation underlies the pathogenesis of hematological malignancies; however, the extent to which deregulated phosphorylation may be involved in B-cell non-Hodgkin lymphoma (B-NHL) pathogenesis is largely unknown. To identify phosphorylation events important in B-NHLs, we performed mass spectrometry-based, label-free, semiquantitative phosphoproteomic profiling of 11 cell lines derived from three B-NHL categories: Burkitt lymphoma, follicular lymphoma, and mantle-cell lymphoma. In all, 6579 unique phosphopeptides, corresponding to 1701 unique phosphorylated proteins, were identified and quantified. The data are available via ProteomeXchange with identifier PXD000658. Hierarchical clustering highlighted distinct phosphoproteomic signatures associated with each lymphoma subtype. Interestingly, germinal center-derived B-NHL cell lines were characterized by phosphorylation of proteins involved in the B-cell receptor signaling. Of these proteins, phosphoprotein associated with glycosphingolipid-enriched microdomains 1 (PAG1) was identified with the most phosphorylated tyrosine peptides in Burkitt lymphoma and follicular lymphoma. PAG1 knockdown resulted in perturbation of the tyrosine phosphosignature of B-cell receptor signaling components. Significantly, PAG1 knockdown increased cell proliferation and response to antigen stimulation of these germinal center-derived B-NHLs. These data provide a detailed annotation of phosphorylated proteins in human lymphoid cancer. Overall, our study revealed the utility of unbiased phosphoproteome interrogation in characterizing signaling networks that may provide insights into pathogenesis mechanisms in B-cell lymphomas.
    American Journal Of Pathology 03/2014; · 4.52 Impact Factor
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    ABSTRACT: Detection of high-frequency BRAF V600E mutations in hairy cell leukemia (HCL) has important diagnostic utility. However, the requisite analytic performance for a clinical assay to routinely detect BRAF V600E mutations in HCL has not been clearly defined. In this study, we sought to determine the level of analytic sensitivity needed for formalin-fixed, paraffin-embedded (FFPE) and frozen samples and to compare the performance of 2 allele-specific polymerase chain reaction (PCR) assays. Twenty-nine cases of classic HCL, including 22 FFPE bone marrow aspirates and 7 frozen specimens from blood or bone marrow were evaluated using a laboratory-developed allele-specific PCR assay and a commercially available allele-specific quantitative PCR assay-myT BRAF Ultra. Also included were 6 HCL variant and 40 non-HCL B-cell lymphomas. Two cases of classic HCL, 1 showing CD5 expression, were truly BRAF V600E-negative based on negative results by PCR and sequencing despite high-level leukemic involvement. Among the remaining 27 specimens, V600E mutations were detected in 88.9% (17/20 FFPE; 7/7 frozen) and 81.5% (15/20 FFPE; 7/7 frozen), for the laboratory-developed and commercial assays, respectively. No mutations were detected among the 46 non-HCL lymphomas. Both assays showed an analytic sensitivity of 0.3% involvement in frozen specimens and 5% in FFPE tissue. On the basis of these results, an assay with high analytic sensitivity is required for the clinical detection of V600E mutations in HCL specimens. Two allele-specific PCR assays performed well in both frozen and FFPE bone marrow aspirates, although detection in FFPE tissue required 5% or more involvement.
    Applied immunohistochemistry & molecular morphology: AIMM / official publication of the Society for Applied Immunohistochemistry 02/2014; · 1.63 Impact Factor
  • A A Sahasrabuddhe, X Chen, F Chung, T Velusamy, M S Lim, K S J Elenitoba-Johnson
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    ABSTRACT: EZH2 (enhancer of zeste homolog 2) is a critical enzymatic subunit of the polycomb repressive complex 2 (PRC2), which trimethylates histone H3 (H3K27) to mediate gene repression. Somatic mutations, overexpression and hyperactivation of EZH2 have been implicated in the pathogenesis of several forms of cancer. In particular, recurrent gain-of-function mutations targeting EZH2 Y641 occur most frequently in follicular lymphoma and aggressive diffuse large B-cell lymphoma and are associated with H3K27me3 hyperactivation, which contributes to lymphoma pathogenesis. However, the post-translational mechanisms of EZH2 regulation are not completely understood. Here we show that EZH2 is a novel interactor and substrate of the SCF E3 ubiquitin ligase β-TrCP (FBXW1). β-TrCP ubiquitinates EZH2 and Jak2-mediated phosphorylation on Y641 directs β-TrCP-mediated EZH2 degradation. RNA interference-mediated silencing of β-TrCP or inhibition of Jak2 results in EZH2 stabilization with attendant increase in H3K27 trimethylation activity. Importantly, the EZH2(Y641) mutants recurrently implicated in lymphoma pathogenesis are unable to bind β-TrCP. Further, endogenous EZH2(Y641) mutants in lymphoma cells exhibit increased EZH2 stability and H3K27me3 hyperactivity. Our studies demonstrate that β-TrCP has an important role in controlling H3K27 trimethylation activity and lymphoma pathogenesis by targeting EZH2 for degradation.Oncogene advance online publication, 27 January 2014; doi:10.1038/onc.2013.571.
    Oncogene 01/2014; · 7.36 Impact Factor
  • Larissa V Furtado, Helmut C Weigelin, Kojo S J Elenitoba-Johnson, Bryan L Betz
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    ABSTRACT: MPL mutation testing is recommended in patients with suspected primary myelofibrosis or essential thrombocythemia who lack the JAK2 V617F mutation. MPL mutations can occur at allelic levels below 15%, which may escape detection by commonly used mutation screening methods such as Sanger sequencing. We developed a novel multiplexed allele-specific PCR assay capable of detecting most recurrent MPL exon 10 mutations associated with primary myelofibrosis and essential thrombocythemia (W515L, W515K, W515A, and S505N) down to a sensitivity of 2.5% mutant allele. Test results were reviewed from 15 reference cases and 1380 consecutive specimens referred to our laboratory for testing. Assay performance was compared to Sanger sequencing across a series of 58 specimens with MPL mutations. Positive cases consisted of 45 with W515L, 6 with S505N, 5 with W515K, 1 with W515A, and 1 with both W515L and S505N. Seven cases had mutations below 5% that were undetected by Sanger sequencing. Ten additional cases had mutation levels between 5% and 15% that were not consistently detected by sequencing. All results were easily interpreted in the allele-specific test. This assay offers a sensitive and reliable solution for MPL mutation testing. Sanger sequencing appears insufficiently sensitive for robust MPL mutation detection. Our data also suggest the relative frequency of S505N mutations may be underestimated, highlighting the necessity for inclusion of this mutation in MPL test platforms.
    The Journal of molecular diagnostics: JMD 08/2013; · 3.48 Impact Factor
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    ABSTRACT: Chromosomal translocations encoding chimeric fusion proteins constitute one of the most common mechanisms underlying oncogenic transformation in human cancer. Fusion peptides resulting from such oncogenic chimeric fusions though unique to specific cancer subtypes are unexplored as cancer biomarkers. Here we show using an approach termed fusion peptide multiple reaction monitoring (FP-MRM) mass spectrometry (MS), the direct identification of different cancer-specific fusion peptides arising from protein chimeras that are generated from juxtaposition of heterologous genes fused by recurrent chromosomal translocations. Using FP-MRM-MS in a clinically relevant scenario, we demonstrate specific, sensitive and unambiguous detection of a specific diagnostic fusion peptide in clinical samples of anaplastic large cell lymphoma (ALCL), but not in a diverse array of benign lymph nodes or other forms of primary malignant lymphomas and cancer-derived cell lines. Our studies highlight the utility of fusion peptides as cancer biomarkers and carry broad implications for the use of protein biomarkers in cancer detection and monitoring.
    Molecular &amp Cellular Proteomics 07/2013; · 7.25 Impact Factor
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    ABSTRACT: Mutations within exon 12 of the JAK2 gene occur in most cases of JAK2 V617F-mutation negative polycythemia vera. Several methods have been developed to identify exon 12 mutations, with both Sanger sequencing and high resolution melting (HRM) being widely used. However, mutations can occur at allelic levels below 15%, which may hamper detection by these methods. We developed a novel fragment analysis-based assay capable of detecting nearly all JAK2 exon 12 mutations associated with polycythemia vera down to a sensitivity of 2% mutant allele. Test results were reviewed from a set of 20 reference cases and 1731 consecutive specimens were referred to our laboratory for testing. Assay performance was compared to sequencing and HRM across a series of 27 specimens with JAK2 exon 12 mutations. Positive cases consisted of 22 with deletion mutations, four with duplications, and one with K539L. Nine cases had mutation levels between 6% and 15% that may not be reliably detected by sequencing or HRM. All cases were easily interpreted in the fragment analysis assay. Sequencing, HRM, and fragment analysis each represent viable platforms for detection of JAK2 exon 12 mutations. Our method performed favorably by providing a simple, robust, and highly sensitive solution for JAK2 exon 12 mutation testing.
    The Journal of molecular diagnostics: JMD 06/2013; · 3.48 Impact Factor
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    ABSTRACT: The mechanisms underlying the pathogenesis of NPM-ALK expressing anaplastic large cell lymphoma (ALCL) are not completely understood. Here we show using an integrated phosphoproteomic and metabolomic strategy that NPM-ALK induces a metabolic shift towards aerobic glycolysis, increased lactate production and biomass production. The metabolic shift is mediated through the ALK phosphorylation of PKM2 at Y105 resulting in decreased enzymatic activity. Small molecule activation of PKM2 or expression of Y105F PKM2 mutant leads to reversal of the metabolic switch with increased oxidative phosphorylation and reduced lactate production coincident with increased cell death, decreased colony formation and reduced tumor growth in an in vivo xenograft model. This study provides comprehensive profiling of the phosphoproteomic and metabolomic consequences of NPM-ALK expression and reveals a novel role of ALK in the regulation of multiple components of cellular metabolism. Our studies show that PKM2 is a novel substrate of ALK and plays a critical role in mediating the metabolic shift towards biomass production and tumorigenesis.
    Blood 06/2013; · 9.06 Impact Factor
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    ABSTRACT: Attachment of ubiquitin to substrate is typically thought to occur via formation of an isopeptide bond between the C-terminal glycine residue of ubiquitin and a lysine residue in the substrate. In vitro, Ube2w is nonreactive with free lysine yet readily ubiquitinates substrate. Ube2w also contains novel residues within its active site that are important for its ability to ubiquitinate substrate. To identify the site of modification we analyzed ubiquitinated substrates by mass spectrometry and found the N-terminal -NH2 group as the site of conjugation. To confirm N-terminal ubiquitination we generated lysine-less and N-terminally blocked versions of one substrate, the polyglutamine disease protein ataxin-3, and showed that Ube2w can ubiquitinate a lysine-less, but not N-terminally blocked ataxin-3. This was confirmed with a second substrate, the neurodegenerative disease protein tau. Finally we directly sequenced the N-terminus of unmodified and ubiquitinated ataxin-3 demonstrating that Ube2w attaches ubiquitin to the N-terminus of its substrates. Together these data demonstrate that Ube2w has novel enzymatic properties that direct ubiquitination of the N-terminus of substrates.
    Journal of Biological Chemistry 05/2013; · 4.65 Impact Factor
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    ABSTRACT: Fragile X-associated tremor ataxia syndrome (FXTAS) results from a CGG repeat expansion in the 50 UTR of FMR1. This repeat is thought to elicit toxicity as RNA, yet disease brains contain ubiqui- tin-positive neuronal inclusions, a pathologic hall- mark of protein-mediated neurodegeneration. We explain this paradox by demonstrating that CGG repeats trigger repeat-associated non-AUG-initiated (RAN) translation of a cryptic polyglycine-containing protein, FMRpolyG. FMRpolyG accumulates in ubiq- uitin-positive inclusions in Drosophila, cell culture, mouse disease models, and FXTAS patient brains. CGG RAN translation occurs in at least two of three possible reading frames at repeat sizes ranging from normal (25) to pathogenic (90), but inclusion formation only occurs with expanded repeats. In Drosophila, CGG repeat toxicity is suppressed by eliminating RAN translation and enhanced by increased polyglycine protein production. These studies expand the growing list of nucleotide repeat disorders in which RAN translation occurs and pro- vide evidence that RAN translation contributes to neurodegeneration.
    Neuron 03/2013; · 15.77 Impact Factor
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    ABSTRACT: Multiple myeloma (MM) represents the malignant proliferation of terminally differentiated B cells, which, in many cases, is associated with the maintenance of high levels of the oncoprotein c-MYC. Overexpression of the histone methyltransferase MMSET (WHSC1/NSD2), due to t(4;14) chromosomal translocation, promotes the proliferation of MM cells along with global changes in chromatin; nevertheless, the precise mechanisms by which MMSET stimulates neoplasia remain incompletely understood. We found that MMSET enhances the proliferation of MM cells by stimulating the expression of c-MYC at the post-transcriptional level. A microRNA (miRNA) profiling experiment in t(4;14) MM cells identified miR-126* as an MMSET-regulated miRNA predicted to target c-MYC mRNA. We show that miR-126* specifically targets the 3'-untranslated region (3'-UTR) of c-MYC, inhibiting its translation and leading to decreased c-MYC protein levels. Moreover, the expression of this miRNA was sufficient to decrease the proliferation rate of t(4;14) MM cells. Chromatin immunoprecipitation analysis showed that MMSET binds to the miR-126* promoter along with the KAP1 corepressor and histone deacetylases, and is associated with heterochromatic modifications, characterized by increased trimethylation of H3K9 and decreased H3 acetylation, leading to miR-126* repression. Collectively, this study shows a novel mechanism that leads to increased c-MYC levels and enhanced proliferation of t(4;14) MM, and potentially other cancers with high MMSET expression.Leukemia advance online publication, 12 October 2012; doi:10.1038/leu.2012.269.
    Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 09/2012; · 10.16 Impact Factor
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    ABSTRACT: Splenic marginal zone lymphoma (SMZL), the most common primary lymphoma of spleen, is poorly understood at the genetic level. In this study, using whole-genome DNA sequencing (WGS) and confirmation by Sanger sequencing, we observed mutations identified in several genes not previously known to be recurrently altered in SMZL. In particular, we identified recurrent somatic gain-of-function mutations in NOTCH2, a gene encoding a protein required for marginal zone B cell development, in 25 of 99 (∼25%) cases of SMZL and in 1 of 19 (∼5%) cases of nonsplenic MZLs. These mutations clustered near the C-terminal proline/glutamate/serine/threonine (PEST)-rich domain, resulting in protein truncation or, rarely, were nonsynonymous substitutions affecting the extracellular heterodimerization domain (HD). NOTCH2 mutations were not present in other B cell lymphomas and leukemias, such as chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL; n = 15), mantle cell lymphoma (MCL; n = 15), low-grade follicular lymphoma (FL; n = 44), hairy cell leukemia (HCL; n = 15), and reactive lymphoid hyperplasia (n = 14). NOTCH2 mutations were associated with adverse clinical outcomes (relapse, histological transformation, and/or death) among SMZL patients (P = 0.002). These results suggest that NOTCH2 mutations play a role in the pathogenesis and progression of SMZL and are associated with a poor prognosis.
    Journal of Experimental Medicine 08/2012; 209(9):1553-65. · 13.21 Impact Factor
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    ABSTRACT: Chronic myeloproliferative neoplasms (MPN) are clonal disorders of hematopoietic stem cells, which fall into distinct categories based on a number of characteristics including the presence of the BCR-ABL1 gene fusion (chronic myelogenous leukemia) or the JAK2(V617F) mutation (polycythemia vera, primary myelofibrosis, and essential thrombocythemia). One of the criteria in the 2008 World Health Organization Classification divides MPN into different categories based on the presence of an underlying genetic abnormality, however the WHO does not currently address the classification of myeloproliferative neoplasms that have more than one genetic abnormality. The coexistence of a JAK2(V617F) mutation and BCR-ABL1 is rare, and to our knowledge, less than 25 cases have been reported in the literature. Our case series examines the clinical, histopathologic, and genetic features of 3 patients with myeloproliferative neoplasms characterized by concomitant BCR-ABL1 and JAK2(V617F). The implications for diagnosis and treatment of patients with concomitant BCR-ABL1 and JAK2(V617F) are discussed as well as how the BCR-ABL1 and JAK2(V617F)-positive clones may be related to one another.
    Diagnostic molecular pathology: the American journal of surgical pathology, part B 07/2012; 21(3):176-83. · 1.58 Impact Factor
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    ABSTRACT: Conjugated polyelectrolyte-antibody hybrid materials promise to enhance the utility of conjugated polymers in bioimaging field. Polymer-antibody conjugates that are biologically safe and highly sensitive and selective to cells are designed to image human T or B lymphocytes. In the clear state, the observed efficiency of luminescence is superior to that of commercially available FITC-antibody probe.
    Advanced Materials 04/2012; 24(18):2479-84. · 14.83 Impact Factor
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    ABSTRACT: Fbxw7α is a member of the F-box family of proteins, which function as the substrate-targeting subunits of SCF (Skp1/Cul1/F-box protein) ubiquitin ligase complexes. Using differential purifications and mass spectrometry, we identified p100, an inhibitor of NF-κB signalling, as an interactor of Fbxw7α. p100 is constitutively targeted in the nucleus for proteasomal degradation by Fbxw7α, which recognizes a conserved motif phosphorylated by GSK3. Efficient activation of non-canonical NF-κB signalling is dependent on the elimination of nuclear p100 through either degradation by Fbxw7α or exclusion by a newly identified nuclear export signal in the carboxy terminus of p100. Expression of a stable p100 mutant, expression of a constitutively nuclear p100 mutant, Fbxw7α silencing or inhibition of GSK3 in multiple myeloma cells with constitutive non-canonical NF-κB activity results in apoptosis both in cell systems and xenotransplant models. Thus, in multiple myeloma, Fbxw7α and GSK3 function as pro-survival factors through the control of p100 degradation.
    Nature Cell Biology 03/2012; 14(4):375-85. · 20.76 Impact Factor
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    ABSTRACT: Anaplastic large cell lymphoma (ALCL) is the most common type of pediatric peripheral T-cell lymphoma. In 70-80% of cases, the chromosomal aberration t(2;5)(p23;q35) results in the juxtaposition of anaplastic lymphoma kinase (ALK) with nucleophosmin (NPM) and the subsequent expression of the NPM-ALK fusion protein. NPM-ALK is a chimeric tyrosine kinase, which induces numerous signaling pathways that drive proliferation and abrogate apoptosis. However, the mechanisms that lead to activation of downstream growth regulatory molecules have not been completely elucidated. Using a mass spectrometry-based phosphoproteomic screen, we identified GSK3β as a signaling mediator of NPM-ALK. Using a selective inhibitor of ALK, we demonstrated that the tyrosine kinase activity of ALK regulates the serine-9 phosphorylation of GSK3β. Expression of NPM-ALK in 293T cells led to an increase of pS(9)-GSK3β (glycogen synthase kinase 3 beta) compared with kinase-defective K210R mutant NPM-ALK, but did not affect total GSK3β levels. Phosphorylation of pS(9)-GSK3β by NPM-ALK was mediated by the PI3K/AKT signaling pathway. ALK inhibition resulted in degradation of GSK3β substrates Mcl-1 and CDC25A, which was recovered upon chemical inhibition of the proteasome (MG132). Furthermore, the degradation of Mcl-1 was recoverable with inhibition of GSK3β. ALK inhibition also resulted in decreased cell viability, which was rescued by GSK3β inhibition. Furthermore, stable knockdown of GSK3β conferred resistance to the growth inhibitory effects of ALK inhibition using viability and colony formation assays. pS(9)-GSK3β and CDC25A were selectively expressed in neoplastic cells of ALK+ALCL tissue biopsies, and showed a significant correlation (P<0.001). Conversely, ALK-ALCL tissue biopsies did not show significant correlation of pS(9)-GSK3β and CDC25A expression (P<0.2). Our results demonstrate that NPM-ALK regulates the phosphorylation of S(9)-GSK3β by PI3K/AKT. The subsequent inhibition of GSK3β activity results in accumulation of CDC25A and Mcl-1, which confers the advantage of growth and protection from apoptosis. These findings provide support for the role of GSK3β as a mediator of NPM-ALK oncogenesis.
    Oncogene 12/2011; 31(32):3733-40. · 7.36 Impact Factor
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    ABSTRACT: In Saccharomyces cerevisae, expanded polyglutamine (polyQ) fragments are assembled into discrete cytosolic aggregates in a process regulated by the molecular chaperones Hsp26, Hsp70, Hsp90, and Hsp104. To better understand how the different chaperones might cooperate during polyQ aggregation, we used sequential immunoprecipitations and mass spectrometry to identify proteins associated with either soluble (Q25) or aggregation-prone (Q103) fragments at both early and later times after induction of their expression. We found that Hsp26, Hsp70, Hsp90, and other chaperones interact with Q103, but not Q25, within the first 2 h. Further, Hsp70 and Hsp90 appear to be partially released from Q103 prior to the maturation of the aggregates and before the recruitment of Hsp104. To test the importance of this seemingly ordered process, we used a chemical probe to artificially enhance Hsp70 binding to Q103. This treatment retained both Hsp70 and Hsp90 on the polyQ fragment and, interestingly, limited subsequent exchange for Hsp26 and Hsp104, resulting in incomplete aggregation. Together, these results suggest that partial release of Hsp70 may be an essential step in the continued processing of expanded polyQ fragments in yeast.
    Journal of Biological Chemistry 11/2011; 286(47):40486-93. · 4.65 Impact Factor
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    ABSTRACT: A number of acute leukemias arise from fusion of the mixed lineage leukemia 1 protein (MLL) N terminus to a variety of fusion partners that have been reported to reside in one or more poorly defined complexes linked to transcription elongation through interactions with the histone H3-K79 methyltransferase DOT1 and positive transcription elongation factor b (P-TEFb). Here we first identify natural complexes (purified through fusion partners AF9, AF4, and ELL) with overlapping components, different elongation activities, and different cofactor associations that suggest dynamic interactions. Then, through reconstitution of defined, functionally active minimal complexes, we identify stable subcomplexes that, through newly defined protein-protein interactions, form distinct higher order complexes. These definitive analyses show, for example, that (i) through direct interactions with AF9 and cyclinT1, family members AF4 and AFF4 independently mediate association of P-TEFb with AF9, (ii) P-TEFb, through direct interactions, provides the link for association of ELL and ELL-associated factors 1 and 2 (EAF1 and EAF2) with AF4, and (iii) in the absence of other factors, DOT1 forms a stable complex with AF9 and does not interact with AF9•AF4•P-TEFb complexes. Finally, we show the importance of defined higher order complex formation in MLL-AF9-mediated transcriptional up-regulation and cell immortalization potential in vivo. Thus, our study provides direct mechanistic insight into the role of fusion partners in MLL fusion-mediated leukemogenesis.
    Proceedings of the National Academy of Sciences 09/2011; 108(38):15751-6. · 9.74 Impact Factor
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    ABSTRACT: Patients with hematologic malignancies not in remission before allogeneic hematopoietic stem cell transplantation (HSCT) have a poor prognosis. To improve the antitumor activity of conditioning, we combined clofarabine with myeloablative doses of busulfan in a phase 1/2 study in nonremission hematologic malignancies. Forty-six patients were enrolled, including 31 patients with nonremission acute myelogenous leukemia (AML). Patients had a median age of 53 years, with a median comorbidity index of 3. Donors were unrelated, HLA mismatched, or both in 59% of patients. Common grade III to IV nonhematologic toxicities included transient transaminitis (50%), mucositis (24%), hand-foot syndrome (13%), transient hypoxia (13%), nausea/vomiting (9%), and diarrhea (9%). All patients engrafted. Complete remission was achieved in 80% of all patients by day +30 and in 100% of AML patients without prior hematopoietic stem cell transplantation. Two-year nonrelapse mortality for all patients was 31%, and overall survival was 28%. In AML, the overall survival was 48% at 1 year and 35% at 2 years. These data suggest that clofarabine combined with myeloablative doses of busulfan is well tolerated, secures engraftment, and possesses significant antitumor activity, particularly in nonremission AML. This study is registered at www.ClinicalTrials.gov under identifier NCT00556452.
    Blood 08/2011; 118(15):4258-64. · 9.06 Impact Factor
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    ABSTRACT: Recurrent fusions of ETS genes are considered driving mutations in a diverse array of cancers, including Ewing's sarcoma, acute myeloid leukemia, and prostate cancer. We investigate the mechanisms by which ETS fusions mediate their effects, and find that the product of the predominant ETS gene fusion, TMPRSS2:ERG, interacts in a DNA-independent manner with the enzyme poly (ADP-ribose) polymerase 1 (PARP1) and the catalytic subunit of DNA protein kinase (DNA-PKcs). ETS gene-mediated transcription and cell invasion require PARP1 and DNA-PKcs expression and activity. Importantly, pharmacological inhibition of PARP1 inhibits ETS-positive, but not ETS-negative, prostate cancer xenograft growth. Finally, overexpression of the TMPRSS2:ERG fusion induces DNA damage, which is potentiated by PARP1 inhibition in a manner similar to that of BRCA1/2 deficiency.
    Cancer cell 05/2011; 19(5):664-78. · 25.29 Impact Factor
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    ABSTRACT: The aim of this study was to better understand how mixed lineage leukemia (MLL) fusion proteins deregulate the expression of genes critical for leukemia. The transforming domain of one of the most common MLL fusion partners, AF9, was immunopurified after expression in myeloblastic M1 cells, and associating proteins were identified by mass spectrometric analysis. Chromatin immunoprecipitation followed by quantitative polymerase chain reaction was used to determine how binding of associating proteins compare across Hoxa9 and Meis1 in cell lines with and without MLL fusion proteins and how binding is altered during gene down-regulation and differentiation. Consistent with earlier purifications of ENL and AF4 from 293 cells, the 90 amino acid C-terminal domain of AF9 associates with many other MLL translocation partners including Enl, Af4, Laf4, Af5q31, Ell, and Af10. This complex, termed elongation assisting proteins (EAPs), also contains the RNA polymerase II C-terminal domain kinase Cdk9/Cyclin T1/T2 (pTEFb) and the histone H3 lysine 79 methyltransferase Dot1L. Myeloid cells transformed by MLL fusions show higher levels and a broader distribution of EAP components at genes critical for leukemia. Inhibition of EAP components pTEFb and Dot1l show that both contribute significantly to activation of Hoxa9 and Meis1 expression. EAP is dynamically associated with the Hoxa9 and Meis1 loci in hematopoietic cells and rapidly dissociates during induction of differentiation. In the presence of MLL fusion proteins, its dissociation is prevented. The findings suggest that MLL fusion proteins deregulate genes critical for leukemia by excessive recruitment and impaired dissociation of EAP from target loci.
    Experimental hematology 01/2011; 39(1):77-86.e1-5. · 3.11 Impact Factor

Publication Stats

2k Citations
620.18 Total Impact Points

Institutions

  • 2009–2014
    • Concordia University–Ann Arbor
      Ann Arbor, Michigan, United States
    • Robert-Bosch Krankenhaus
      Stuttgart, Baden-Württemberg, Germany
  • 2008–2013
    • University of Michigan
      • Department of Pathology
      Ann Arbor, Michigan, United States
    • University of Washington Seattle
      • Department of Pathology
      Seattle, WA, United States
  • 1998–2009
    • University of Utah
      • • Department of Pathology
      • • Division of Rheumatology
      • • ARUP Institute for Clinical and Experimental Pathology
      Salt Lake City, UT, United States
    • National Human Genome Research Institute
      Maryland, United States
  • 2007
    • Pennsylvania State University
      • Department of Pathology
      University Park, MD, United States
    • University of Texas MD Anderson Cancer Center
      • Department of Hematopathology
      Houston, TX, United States
  • 2006
    • ARUP Laboratories: A National Reference Laboratory
      Salt Lake City, Utah, United States
  • 2000
    • Arup
      Londinium, England, United Kingdom
  • 1997–1998
    • National Institutes of Health
      • Laboratory of Pathology
      Bethesda, MD, United States
    • National Cancer Institute (USA)
      • Laboratory of Pathology
      Maryland, United States
  • 1996
    • Rhode Island Hospital
      Providence, Rhode Island, United States
  • 1995
    • Roger Williams University
      Bristol, Rhode Island, United States