The AF4·MLL fusion protein is capable of inducing ALL in mice without requirement of MLL·AF4

Blood ◽  
2010 ◽  
Vol 115 (17) ◽  
pp. 3570-3579 ◽  
Author(s):  
Adelheid Bursen ◽  
Karen Schwabe ◽  
Brigitte Rüster ◽  
Reinhard Henschler ◽  
Martin Ruthardt ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Fusion Protein ◽  
Fusion Proteins ◽  
Lymphoblastic Leukemia ◽  
Chromosomal Translocation ◽  
Disease Development ◽  
Fusion Genes ◽  
Genetic Aberration ◽  
Biphenotypic Acute Leukemia ◽  
Observation Period

Abstract The chromosomal translocation t(4;11)(q21;q23) is the most frequent genetic aberration of the human MLL gene, resulting in high-risk acute lymphoblastic leukemia (ALL). To elucidate the leukemogenic potential of the fusion proteins MLL·AF4 and AF4·MLL, Lin−/Sca1+ purified cells (LSPCs) were retrovirally transduced with either both fusion genes or with MLL·AF4 or AF4·MLL alone. Recipients of AF4·MLL- or double-transduced LSPCs developed pro-B ALL, B/T biphenotypic acute leukemia, or mixed lineage leukemia. Transplantation of MLL·AF4- or mock-transduced LSPCs did not result in disease development during an observation period of 13 months. These findings indicate that the expression of the AF4·MLL fusion protein is capable of inducing acute lymphoblastic leukemia even in the absence of the MLL·AF4 fusion protein. In view of recent findings, these results may imply that t(4;11) leukemia is based on 2 oncoproteins, providing an explanation for the very early onset of disease in humans.

scholarly journals Biology of t(6;11) Fusion Proteins and Their Role in MLL-Rearranged Acute Leukemia Lineage Determination

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5033-5033
Author(s):  
Arpita Kundu ◽  
Eric Kowarz ◽  
Jennifer Reis ◽  
Rolf Marschalek
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Acute Leukemia ◽  
Fusion Proteins ◽  
Lymphoblastic Leukemia ◽  
Chromosomal Translocation ◽  
Fusion Genes ◽  
Disease Phenotype ◽  
Acute Leukemias

Chromosomal translocations are genetic rearrangements where a chromosomal segment is transferred to a non-homologous chromosome which give rise to novel chimeras. Chromosomal rearrangements play a significant role in the development of acute leukemias (acute lymphoblastic leukemia (ALL) or acute myeloid leukemia (AML)). Chromosomal translocation events occurring at 11q23 involving the KMT2A or Mixed-Lineage Leukemia (MLL) gene (n=102) can be diagnosed in about 5-10% of all acute leukemia patients (Marschalek Ann Lab Med 2016), especially prevalent in infant acute leukemias (up to 70% of cases). Different chromosomal translocation partner genes (such as AF4, AF6, AF9orENL and ELL) account for the majority of leukemia cases and have their genomic breakpoints within a major breakpoint cluster region (BCR intron 9-11; Meyer et. al. Leukemia 2018). Some rearrangements are specifically associated with particular disease phenotype e.g. the majority of ALL patients (~ 90%) are mainly caused by the following gene fusions, MLL-AF4, MLL-AF9, MLL-ENL. We are interested in a rare but yet drastic chromosomal translocation t(6;11)(q27;q23) which fuses KMT2A/MLL to Afadin (AFDN/AF6) gene. This chromosomal rearrangement has a very poor prognosis (survival-rate is ~10%) and is predominantly diagnosed in patients with high-risk AML. In this project, we investigate the molecular consequences of two different MLL-AF6 fusions and their corresponding reciprocal AF6-MLL fusions. MLL-AF6 fusions are mainly occurring within MLL intron 9 to 11 and are associated with an AML disease phenotype, while the same fusion occurring within the minor breakpoints region in MLL intron 21 until exon (ex) 24 are mainly diagnosed with T-ALL (T-cell acute lymphoblastic leukemia) disease phenotype. The molecular mechanism that determines the resulting disease phenotype is yet unknown. Therefore, we cloned all of these t(6;11) fusion proteins in order to investigate the functional consequences of the two different breakpoints (MLLex1-9::AF6ex2-30, AF6ex1::MLLex10-37; MLLex1-21::AF6ex2-30, AF6ex1::MLLex22-37). All 4 fusion genes were introduced into our inducible Sleeping Beauty system (Ivics et. al. Mobile DNA 2010; Kowarz et. al. Biotechnol J. 2015) and stably transfected reporter cell lines. Basically, these 4 fusion proteins differ only in the presence or absence of their Plant homeodomain 1-3/Bromodomain (PHD1-3/BD) domain (see Figure 1). The PHD domain regulates the epigenetic and transcriptional regulatory functions of wildtype MLL. Subsequently, we analyzed gene expression differences by the MACE-Seq (Massive Analyses of cDNA Ends). MACE data revealed fundamental differences in gene expression profiles when analyzing the two different sets of t(6;11) fusion genes. The resulting profiles have similarities to either AML or T-ALL and might give a rational explanation for the different lineages in these t(6;11) patients. Altogether, these results notably indicate that our study will provide a novel insight into this type of high-risk leukemia and subsequently will be useful for developing of novel and appropriate therapeutic strategies against acute leukemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals PAX5-ELN oncoprotein promotes multistep B-cell acute lymphoblastic leukemia in mice

2018 ◽  
Vol 115 (41) ◽  
pp. 10357-10362 ◽  
Author(s):  
Laura Jamrog ◽  
Guillaume Chemin ◽  
Vincent Fregona ◽  
Lucie Coster ◽  
Marlène Pasquet ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Fusion Proteins ◽  
Lymphoblastic Leukemia ◽  
Chromosomal Translocation ◽  
Suppressor Gene ◽  
In Vivo Model ◽  
Functional Studies

PAX5 is a well-known haploinsufficient tumor suppressor gene in human B-cell precursor acute lymphoblastic leukemia (B-ALL) and is involved in various chromosomal translocations that fuse a part of PAX5 with other partners. However, the role of PAX5 fusion proteins in B-ALL initiation and transformation is ill-known. We previously reported a new recurrent t(7;9)(q11;p13) chromosomal translocation in human B-ALL that juxtaposed PAX5 to the coding sequence of elastin (ELN). To study the function of the resulting PAX5-ELN fusion protein in B-ALL development, we generated a knockin mouse model in which the PAX5-ELN transgene is expressed specifically in B cells. PAX5-ELN–expressing mice efficiently developed B-ALL with an incidence of 80%. Leukemic transformation was associated with recurrent secondary mutations on Ptpn11, Kras, Pax5, and Jak3 genes affecting key signaling pathways required for cell proliferation. Our functional studies demonstrate that PAX5-ELN affected B-cell development in vitro and in vivo featuring an aberrant expansion of the pro-B cell compartment at the preleukemic stage. Finally, our molecular and computational approaches identified PAX5-ELN–regulated gene candidates that establish the molecular bases of the preleukemic state to drive B-ALL initiation. Hence, our study provides a new in vivo model of human B-ALL and strongly implicates PAX5 fusion proteins as potent oncoproteins in leukemia development.

Interaction of AF4 Wildtype and AF4•MLL Fusion Protein with SIAH Proteins: Indication for T(4;11) Pathobiology?.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4268-4268
Author(s):  
Adelheid Bursen ◽  
Moritz Sven ◽  
Gaussmann Anne ◽  
Dingermann Theo ◽  
Marschalek Rolf
Keyword(s):  
Early Childhood ◽  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Fusion Protein ◽  
Lymphoblastic Leukemia ◽  
Recombination Process ◽  
Fusion Genes ◽  
Terminal Portion ◽  
Mll Gene ◽  
Proteosomal Degradation

Abstract The human AF4 gene (4q21) is recurrently involved in recipro cal translocations to the MLL gene (11q23), correlated with high-risk acute lymphoblastic leukemia (ALL) in infants and early childhood. The t(4;11) translocation is one of the most frequent MLL translocations known today. In general, MLL translocations are the result of an illegitimate recombination process leading to reciprocal fusions of unrelated translocation partner (TP) genes with the MLL gene. Due to the constant presence of the derivative(11) product, the hypothesis was posed that only MLL•TP fusion genes are responsible for the leukemogenic process. This concept has been successfully tested for some known MLL fusions, while some other MLL fusions failed. Here, we demonstrate growth transforming potential of AF4 wildtype and the AF4•MLL fusion protein. The underlying oncogenic mechanism involves the two E3 ubiquitin ligases SIAH1 and SIAH2, the N-terminal portion of AF4 and the protection of the AF4•MLL fusion protein against proteosomal degradation. Supported by grant Ma 1786/4-1 from the DFG.

scholarly journals The Structural Characterization of Tumor Fusion Genes and Proteins

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Dandan Wang ◽  
Daixi Li ◽  
Guangrong Qin ◽  
Wen Zhang ◽  
Jian Ouyang ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Fusion Proteins ◽  
Chromosomal Translocation ◽  
Structural Features ◽  
Fusion Partner ◽  
Structural Level ◽  
Fusion Genes ◽  
And Function ◽  
Disordered Regions

Chromosomal translocation, which generates fusion proteins in blood tumor or solid tumor, is considered as one of the major causes leading to cancer. Recent studies suggested that the disordered fragments in a fusion protein might contribute to its carcinogenicity. Here, we investigated the sequence feature near the breakpoints in the fusion partner genes, the structure features of breakpoints in fusion proteins, and the posttranslational modification preference in the fusion proteins. Results show that the breakpoints in the fusion partner genes have both sequence preference and structural preference. At the sequence level, nucleotide combination AG is preferred before the breakpoint and GG is preferred at the breakpoint. At the structural level, the breakpoints in the fusion proteins prefer to be located in the disordered regions. Further analysis suggests the phosphorylation sites at serine, threonine, and the methylation sites at arginine are enriched in disordered regions of the fusion proteins. Using EML4-ALK as an example, we further explained how the fusion protein leads to the protein disorder and contributes to its carcinogenicity. The sequence and structural features of the fusion proteins may help the scientific community to predict novel breakpoints in fusion genes and better understand the structure and function of fusion proteins.

scholarly journals Activity of immunoproteasome inhibitor ONX-0914 in acute lymphoblastic leukemia expressing MLL–AF4 fusion protein

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tyler W. Jenkins ◽  
Sondra L. Downey-Kopyscinski ◽  
Jennifer L. Fields ◽  
Gilbert J. Rahme ◽  
William C. Colley ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Fusion Protein ◽  
Active Sites ◽  
Lymphoblastic Leukemia ◽  
Clinical Stage ◽  
Proteasome Inhibitors ◽  
Chromosomal Translocation ◽  
Lymphoid Tissues ◽  
Synergistic Cytotoxicity ◽  
Mantle Cell

AbstractProteasome inhibitors bortezomib and carfilzomib are approved for the treatment of multiple myeloma and mantle cell lymphoma and have demonstrated clinical efficacy for the treatment of acute lymphoblastic leukemia (ALL). The t(4;11)(q21;q23) chromosomal translocation that leads to the expression of MLL–AF4 fusion protein and confers a poor prognosis, is the major cause of infant ALL. This translocation sensitizes tumor cells to proteasome inhibitors, but toxicities of bortezomib and carfilzomib may limit their use in pediatric patients. Many of these toxicities are caused by on-target inhibition of proteasomes in non-lymphoid tissues (e.g., heart muscle, gut, testicles). We found that MLL–AF4 cells express high levels of lymphoid tissue-specific immunoproteasomes and are sensitive to pharmacologically relevant concentrations of specific immunoproteasome inhibitor ONX-0914, even in the presence of stromal cells. Inhibition of multiple active sites of the immunoproteasomes was required to achieve cytotoxicity against ALL. ONX-0914, an inhibitor of LMP7 (ß5i) and LMP2 (ß1i) sites of the immunoproteasome, and LU-102, inhibitor of proteasome ß2 sites, exhibited synergistic cytotoxicity. Treatment with ONX-0914 significantly delayed the growth of orthotopic ALL xenograft tumors in mice. T-cell ALL lines were also sensitive to pharmacologically relevant concentrations of ONX-0914. This study provides a strong rationale for testing clinical stage immunoproteasome inhibitors KZ-616 and M3258 in ALL.

scholarly journals NUP214 in Leukemia: It’s More than Transport

Cells ◽  
2019 ◽  
Vol 8 (1) ◽  
pp. 76 ◽  
Author(s):  
Adélia Mendes ◽  
Birthe Fahrenkrog
Keyword(s):  
Fusion Protein ◽  
Transcription Regulation ◽  
Chromatin Remodeling ◽  
Nuclear Export ◽  
Fusion Proteins ◽  
Lymphoblastic Leukemia ◽  
Chromosomal Translocation ◽  
Nuclear Pore ◽  
Cellular Effects ◽  
Sequestosome 1

NUP214 is a component of the nuclear pore complex (NPC) with a key role in protein and mRNA nuclear export. Chromosomal translocations involving the NUP214 locus are recurrent in acute leukemia and frequently fuse the C-terminal region of NUP214 with SET and DEK, two chromatin remodeling proteins with roles in transcription regulation. SET-NUP214 and DEK-NUP214 fusion proteins disrupt protein nuclear export by inhibition of the nuclear export receptor CRM1, which results in the aberrant accumulation of CRM1 protein cargoes in the nucleus. SET-NUP214 is primarily associated with acute lymphoblastic leukemia (ALL), whereas DEK-NUP214 exclusively results in acute myeloid leukemia (AML), indicating different leukemogenic driver mechanisms. Secondary mutations in leukemic blasts may contribute to the different leukemia outcomes. Additional layers of complexity arise from the respective functions of SET and DEK in transcription regulation and chromatin remodeling, which may drive malignant hematopoietic transformation more towards ALL or AML. Another, less frequent fusion protein involving the C terminus of NUP214 results in the sequestosome-1 (SQSTM1)-NUP214 chimera, which was detected in ALL. SQSTM1 is a ubiquitin-binding protein required for proper autophagy induction, linking the NUP214 fusion protein to yet another cellular mechanism. The scope of this review is to summarize the general features of NUP214-related leukemia and discuss how distinct chromosomal translocation partners can influence the cellular effects of NUP214 fusion proteins in leukemia.

scholarly journals E2A-PBX1 functions as a coactivator for RUNX1 in acute lymphoblastic leukemia

Blood ◽  
2020 ◽  
Vol 136 (1) ◽  
pp. 11-23 ◽  
Author(s):  
Wen-Chieh Pi ◽  
Jun Wang ◽  
Miho Shimada ◽  
Jia-Wei Lin ◽  
Huimin Geng ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Cell Fate ◽  
Cell Transformation ◽  
Lymphoblastic Leukemia ◽  
Gene Activation ◽  
Chromosomal Translocation ◽  
Direct Interaction ◽  
Binding Activity ◽  
Specific Cell ◽  
Dna Binding Activity

Abstract E2A, a basic helix-loop-helix transcription factor, plays a crucial role in determining tissue-specific cell fate, including differentiation of B-cell lineages. In 5% of childhood acute lymphoblastic leukemia (ALL), the t(1,19) chromosomal translocation specifically targets the E2A gene and produces an oncogenic E2A-PBX1 fusion protein. Although previous studies have shown the oncogenic functions of E2A-PBX1 in cell and animal models, the E2A-PBX1–enforced cistrome, the E2A-PBX1 interactome, and related mechanisms underlying leukemogenesis remain unclear. Here, by unbiased genomic profiling approaches, we identify the direct target sites of E2A-PBX1 in t(1,19)–positive pre-B ALL cells and show that, compared with normal E2A, E2A-PBX1 preferentially binds to a subset of gene loci cobound by RUNX1 and gene-activating machineries (p300, MED1, and H3K27 acetylation). Using biochemical analyses, we further document a direct interaction of E2A-PBX1, through a region spanning the PBX1 homeodomain, with RUNX1. Our results also show that E2A-PBX1 binding to gene enhancers is dependent on the RUNX1 interaction but not the DNA-binding activity harbored within the PBX1 homeodomain of E2A-PBX1. Transcriptome analyses and cell transformation assays further establish a significant RUNX1 requirement for E2A-PBX1–mediated target gene activation and leukemogenesis. Notably, the RUNX1 locus itself is also directly activated by E2A-PBX1, indicating a multilayered interplay between E2A-PBX1 and RUNX1. Collectively, our study provides the first unbiased profiling of the E2A-PBX1 cistrome in pre-B ALL cells and reveals a previously unappreciated pathway in which E2A-PBX1 acts in concert with RUNX1 to enforce transcriptome alterations for the development of pre-B ALL.

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