Characterization of AK2 Gene Function in Zebrafish Hematopoiesis

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2185-2185
Author(s):  
Alberto F Rissone ◽  
Guridevi Jayashree Jagadeesh ◽  
Karen Simon ◽  
Kevin Bishop ◽  
Milton English ◽  
...  
Keyword(s):  
Embryo Development ◽  
Molecular Mechanisms ◽  
Conflicts Of Interest ◽  
Expression Patterns ◽  
Model Systems ◽  
Published Data ◽  
Hematopoietic System ◽  
Alternatively Spliced ◽  
Isoform B

Abstract Abstract 2185 Objective: The Adenylate Kinase (AK) gene family consists of 7 different members (AK1-7) that contribute to energy metabolism of the cells by converting ATP (or GTP) and free AMP to ADP (or GDP) and free ADP. AKs are critical players in ensuring cellular energy homeostasis in all tissues and are generally involved in a broad range of cellular functions. Among AKs, AK2 is uniquely located in the mitochondrial intermembrane space and has been implicated in Caspase 10-mediated apoptosis, although the published data remain controversial. More recently, it was demonstrated that mutations of the AK2 gene cause reticular dysgenesis, an autosomal recessive form of severe combined immunodeficiency (SCID). Reticular dysgenesis is characterized by an early differentiation arrest in the granulocyte lineage and impaired lymphoid maturation resulting in overwhelming infections and high lethality in affected patients. Moreover, patients commonly present with bilateral sensorineural deafness. The mechanisms underlying the biological consequences of AK2-defieincy remain unclear and the generation and characterization study of model systems is expected to provide useful insigths. Ak2 gene-targeted mice have not been reported in the literature, but lines carrying homozygous inactivating retroviral insertions have been shown to be embryonically lethal (our unpublished observations). Because of the known advantages of zebrafish as model system for developmental studies and the similarities of hematopoiesis in zebrafish and higher vertebrates, we set out to investigate the function of the zebrafish ak2 gene in development, with particular emphasis on hematopoiesis. Results: Similar to humans, we found that two different alternatively spliced isoforms of the ak2 gene (Isoform A and Isoform B) are expressed in zebrafish. By Real-Time PCR and In situ Hybridization (ISH) we analyzed the expression of both ak2 isoforms during embryo development. Preliminary data indicate that Isoform A is more abundantly represented than Isoform B during embryo development. ISH analysis showed that the two isoforms have different spatial expression patterns. These data suggest different functionalities for ak2 isoforms during embryo development. To explore such hypothesis, we injected two different morpholinos (MOs) targeting the ak2 isoforms. Downregulation of both ak2 isoforms phenocopied the human disease and resulted in a strong reduction of developing lymphocytes. Moreover we observed a hypochromic phenotype that also suggested impairment of the erythroid lineage. ISH experiments are underway to better define the affected hematopoietic lineages. Interestingly, ak2 MOs-injected embryos showed also developmental defects beyond the hematopoietic system, such as abnormal jaw development. Future studies will focus on the characterization of the specific function of the alternatively spliced ak2 isoforms. Conclusions: We show that the transcription features of the AK2 gene are conserved in zebrafish. The observed differential expression patterns of the zebrafish ak2 isoforms may provide new insights into the function of AK2 in the development of the hematopoietic system, as well as other organs and offers prospects for the understanding of the molecular mechanisms involved in reticular dysgenesis. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Molecular Characterization of Photoperiodic Flowering in Cultivated Strawberry

HortScience ◽  
2005 ◽  
Vol 40 (4) ◽  
pp. 1105D-1105
Author(s):  
Philip Stewart ◽  
Daniel Sargent ◽  
Thomas Davis ◽  
Kevin Folta
Keyword(s):  
Arabidopsis Thaliana ◽  
Molecular Characterization ◽  
Molecular Mechanisms ◽  
Expression Patterns ◽  
Model Systems ◽  
Day Length ◽  
Photoperiodic Flowering ◽  
Long Day ◽  
Flowering Control

The molecular mechanisms governing photoperiodic flowering has been well defined in the model systems of Arabidopsis thaliana(a facultative long-day plant) and rice (a short-day plant). Photoperiodic flowering control is of great interest to strawberry (Fragaria×ananassa) breeders and growers, and the genetics of photoperiodic flowering have been well studied, indicating that response to day-length is regulated by a small number of genetic loci. Cultivated strawberry is octoploid, so identification of these loci through forward genetic analyses is not practical. Since the componentry of the flowering response is generally conserved between monocots and dicots, we may assume that similar, if not identical, systems are functioning in strawberry as well. The goal of this work is to understand how cultivars likely containing identical photoperiod-sensing components are differentially sensitive to daylength. The expression patterns of genes relevant to the floraltransition were assessed under specific photoperiod conditions to assess similarities and/or differences to the model systems.

Focal Adhesion Kinase Inactivation Reduces the Development of Acute Leukemia and Partially Rescues Hematopoietic Stem Cell Defects in Pten-Knockout Mice

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 864-864
Author(s):  
Dewen You ◽  
Andrew Volk ◽  
Clare Sun ◽  
Junping Xin ◽  
Geunhyoung Ha ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Phosphatase Activity ◽  
Focal Adhesion ◽  
Protein Phosphatase ◽  
Molecular Mechanisms ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Hematopoietic Stem ◽  
Hematopoietic System ◽  
Polycytidylic Acid

Abstract Abstract 864 Phosphatase and tensin homolog on chromosome 10 (Pten) is a tumor suppressor which possesses both lipid and protein phosphatase activities. Mutations and epigenetic inactivations of the Pten gene are commonly detected in a large number of tissue malignancies, including leukemias and lymphomas. Studies using Hematopoietic Pten-knockout in adult mice (Pten−/−) have demonstrated that Pten plays a critical role in maintaining the homeostasis of bone marrow (BM) hematopoiesis. Pten inactivation promotes the proliferation and peripheral mobilization of BM hematopoietic stem cells (HSCs). Pten−/− mice develop myeloproliferative disorders (MPD) within days, followed by acute leukemic transformation. Most previous studies attributed such phenotypic changes observed in Pten−/− mice to excessive activation of the PI3K/AKT/mTOR signal, a consequence of the loss of Pten's lipid phosphatase activity. However, the role of Pten's protein phosphatase activity in the regulation of HSCs and leukemogenesis is not well studied. Focal adhesion kinase (Fak) is a critical substrate for the protein phosphatase activity of Pten. Dysregulation of Fak has been observed in many cancers, including acute myeloid leukemias (AML) and acute lymphocytic leukemias (ALL). Therefore, we postulated that Fak might play a pivotal role in the development and progression of leukemia following Pten deletion. To test this hypothesis, we generated Mx1-Cre+Ptenfl/flFakfl/fl mice (an interferon-inducible Pten and Fak compound-knockout, Pten−/−Fak−/−) in which both the Pten and Fak genes in the hematopoietic system are deleted upon injection of polyinosinic-polycytidylic acid (pI-pC). Our results showed that the genetic inactivation of Fak can partially rescue HSC defects associated with Pten deficiency. We found that peripheral mobilization of HSCs in Pten−/−Fak−/− mice is significantly reduced compared to Pten−/− mice. As a consequence, more long-term HSCs (LT-HSCs) are preserved in the BM of Pten−/−Fak−/− mice compared to Pten−/− mice. Transplantation studies suggested that the hematopoietic reconstitutive capacity of Pten−/−Fak−/− HSCs is significantly improved compared to Pten−/− HSCs. Although Fak deletion fails to prevent the development of MPD in Pten−/− mice, Fak deletion does significantly reduce the frequency of AML/ALL, also significantly delays the onset of AML/ALL in comparison to Pten−/− mice. This study suggests that Fak might be a potential target for preventing the MPD-to-AML/ALL transformation and therefore blocking the Fak activity may hold a promise for a novel anti-leukemia therapy. The molecular mechanisms underlying the phenotype restoration of Pten−/− mice by Fak deletion in the hematopoietic system are actively being studied in our laboratory. Disclosures: No relevant conflicts of interest to declare.

The O-Linked N-Acetylglucosamine Transferase Modifies HOXA9 and Inhibits HOXA9-Driven Immortalization

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4901-4901
Author(s):  
Jean-Francois M Rual ◽  
Jay L. Hess ◽  
Tao Xu ◽  
Cailin Collins ◽  
Honglai Zhang ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Bone Marrow Cells ◽  
Conflicts Of Interest ◽  
Therapeutic Benefit ◽  
Core Element ◽  
Hematopoietic Stem ◽  
Primary Bone ◽  
Functional Relevance ◽  
Primary Bone Marrow

Abstract The homeodomain-containing transcription factor HOXA9 is a core element of the HOXA9 enhanceosome, a critical DNA-protein complex that regulates hematopoietic stem cell self-renewal during hematopoiesis. Several genetic mutations observed in acute myeloid leukemia (AML) patients, including MLL translocations, are associated with aberrant up regulation of HOXA9, thus disrupting the hematopoietic balance towards leukemogenesis. While analyses of HOXA9 and cofactors have uncovered fundamental aspects of the mechanisms through which these proteins mediate their functions, questions remain. For example, what molecular mechanisms contribute to switching HOXA9 enhanceosomes off during myeloid differentiation? Could these mechanisms be targeted for the therapeutic benefit of leukemia patients? Characterization of the molecular interactions in which HOXA9 enhanceosome proteins are involved should shed light on the mechanisms that govern these proteins during both normal hematopoiesis and leukemogenesis. We recently discovered that HOXA9 interacts physically with OGT, the only O-linked N-acetyl glucosamine transferase in humans. We also demonstrated that HOXA9 is O-GlcNAcylated by OGT. Investigation of the functional relevance of this interaction to HOXA9-driven leukemogenesis is currently under way using interaction- and O-GlcNAcylation-deficient alleles of HOXA9 in a colony formation assay. Our preliminary results suggest that OGT inhibits HOXA9’s ability to transform primary bone marrow cells, thus suggesting OGT is a potential tumor suppressor of HOXA9-driven leukemogenesis. Current efforts focus on further dissecting the molecular interplay occurring between HOXA9 and OGT on chromatin, its impact on the regulation of HOXA9 targets and its role in HOXA9-driven leukemogenesis. Work is also under way to identify factors involved in the OGT-mediated regulation of HOXA9 enhanceosomes. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Comprehensive Analysis of Ubiquitously Expressed Genes in Human, From a Data-Driven Perspective

2021 ◽  
Author(s):  
Jianlei Gu ◽  
Jiawei Dai ◽  
Hui Lu ◽  
Hongyu Zhao
Keyword(s):  
Beta Cells ◽  
Molecular Mechanisms ◽  
Expression Patterns ◽  
Data Driven ◽  
Gene Clustering ◽  
Internal Reference ◽  
Temporal Gene Expression ◽  
Human Genes ◽  
Extensive Collection

AbstractComprehensive characterization of spatial and temporal gene expression patterns in humans is critical for uncovering regulatory codes of the human genome and understanding molecular mechanisms of human diseases. The ubiquitously expressed genes (UEGs) refer to those genes expressed across a majority, if not all, phenotypic and physiological conditions of an organism. It is known that many human genes are broadly expressed across tissues. However, most previous UEG studies have only focused on providing a list of UEGs without capturing their global expression patterns, thus limiting the potential use of UEG information. In this article, we propose a novel data-driven framework to leverage the extensive collection of ∼40,000 human transcriptomes to derive a list of UEGs and their corresponding global expression patterns, which offers a valuable resource to further validate and characterize human UEGs. Our results suggest that about half (12,234; 49.01%) of the human genes are expressed in at least 80% of human transcriptomes and the median size of the human transcriptome is 16,342 (65.44%). This suggests that the average difference in gene content between human transcriptomes is only 16.43%. Through gene clustering, we identified a set of UEGs, named LoVarUEGs, that have stable expression across human transcriptomes and can be used as internal reference genes for expression measurement. To further demonstrate the usefulness of this resource, we evaluated the uniqueness of repression for 16 previously predicted disallowed genes in islets beta cells and found that seven of these genes showed relatively more varied expression patterns, suggesting that the repression of these genes may not be unique to islets beta cells. We have made our resource publicly available at https://github.com/macroant/HumanUEGs.

scholarly journals Genome-Wide Identification and Characterization of bHLH Transcription Factors Related to Anthocyanin Biosynthesis in Red Walnut (Juglans regia L.)

2021 ◽  
Vol 12 ◽  
Author(s):  
Wei Zhao ◽  
Yonghui Liu ◽  
Lin Li ◽  
Haijun Meng ◽  
Ying Yang ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Gene Family ◽  
Developmental Stage ◽  
Molecular Mechanisms ◽  
Anthocyanin Biosynthesis ◽  
Expression Profiles ◽  
Juglans Regia ◽  
Expression Patterns ◽  
Regulatory Elements

Basic helix-loop-helix (bHLH) proteins are transcription factors (TFs) that have been shown to regulate anthocyanin biosynthesis in many plant species. However, the bHLH gene family in walnut (Juglans regia L.) has not yet been reported. In this study, 102 bHLH genes were identified in the walnut genome and were classified into 15 subfamilies according to sequence similarity and phylogenetic relationships. The gene structure, conserved domains, and chromosome location of the genes were analyzed by bioinformatic methods. Gene duplication analyses revealed that 42 JrbHLHs were involved in the expansion of the walnut bHLH gene family. We also characterized cis-regulatory elements of these genes and performed Gene Ontology enrichment analysis of gene functions, and examined protein-protein interactions. Four candidate genes (JrEGL1a, JrEGL1b, JrbHLHA1, and JrbHLHA2) were found to have high homology to genes encoding bHLH TFs involved in anthocyanin biosynthesis in other plants. RNA sequencing revealed tissue- and developmental stage-specific expression profiles and distinct expression patterns of JrbHLHs according to phenotype (red vs. green leaves) and developmental stage in red walnut hybrid progeny, which were confirmed by quantitative real-time PCR analysis. All four of the candidate JrbHLH proteins localized to the nucleus, consistent with a TF function. These results provide a basis for the functional characterization of bHLH genes and investigations on the molecular mechanisms of anthocyanin biosynthesis in red walnut.

Csf1r Is a Downstream Target of C/EBPβ in Ly6C¯ Monocytes

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 994-994
Author(s):  
Akihiro Tamura ◽  
Hideyo Hirai ◽  
Asumi Yokota ◽  
Atsushi Sato ◽  
Tsukimi Shoji ◽  
...  
Keyword(s):  
Binding Sites ◽  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Conflicts Of Interest ◽  
Expression Patterns ◽  
Hek293 Cells ◽  
Hematopoietic Stem ◽  
Downstream Target ◽  
Inflammatory Monocytes ◽  
Drastic Increase

Abstract Currently, monocytes are classified into at least two subsets. Classical monocytes, also known as inflammatory monocytes (a Ly6C+ subset in mice and a CD14+ CD16− subset in human), are involved in innate immune responses. On the other hand, patrolling monocytes (a Ly6C− subset in mice and a CD14− CD16+ subset in human) have been recently identified. Ly6C− monocytes are found attached on the luminal side of endothelium and scavenge microparticles. Developmentally, Ly6C+ and Ly6C− monocytes share common monocyte progenitors (cMoPs), or Ly6C− monocytes might be converted from Ly6C+ monocytes. Although involvement of Ly6C− monocytes in various kinds of diseases has been reported, molecular mechanisms which regulate the homeostasis of Ly6C− monocytes are largely unknown. CCAAT/Enhancer Binding Protein β (C/EBPβ) is a leucine zipper type transcription factor. We and others have previously shown that C/EBPβ is required for stress-induced granulopoiesis (Hirai et al. Nat Immunol, 2006, Satake et al. J Immunol, 2012, Hayashi et al. Leukemia 2013). However, its roles in steady state hematopoiesis remain relatively unknown. We have recently found that peripheral blood monocytes are significantly reduced in Cebpb−/− mice (Tamura et al. Biochem Biophys Res Commun, 2015). In addition, last year in this meeting, we have reported that Cebpb mRNA is highly upregulated during differentiation from myeloid progenitors or Ly6C+ monocytes to Ly6C− monocytes, and that Ly6C− monocytes are almost completely absent in Cebpb−/− mice due to enhanced cell death [Abstract #224]. Here, we further investigated the molecular mechanisms underlying C/EBPβ-dependent survival of Ly6C− monocytes. In this study, we focused on the regulation of Csf1r (also known as M-CSF receptor). Csf1r is an essential molecule for the development and survival of monocytes. To determine the developmental stages at which Csf1r plays critical roles, we measured the expressions of Csf1r mRNA in hematopoietic stem/progenitor cells and monocyte subsets obtained from wild-type (WT) mice. Csf1r mRNA was expressed at at low levels in hematopoietic stem/progenitors including macrophage dendritic precursors (MDPs) and cMoPs. Csf1r mRNA started to be upregulated in Ly6C+ monocytes, followed by a drastic increase in Ly6C− monocytes. These expression patterns were quite similar to those of Cebpb, suggesting the close relationship between Csf1r and C/EBPβ. Interestingly, such drastic increase of Csf1r mRNA in Ly6C− monocytes was blunted in Cebpb−/− mice, and protein levels of Csf1r in Cebpb−/− Ly6C− monocytes were significantly lower than those in WT Ly6C− monocytes. In order to evaluate the effect of C/EBPβ overexpression on Csf1r expression, EML cells, a mouse hematopoietic stem cell line, were engineered to express C/EBPβ-estrogen receptor (ER) fusion protein or ER alone. Nuclear translocation of C/EBPβ-ER in the presence of tamoxifen resulted in significantly increased levels of Csf1r mRNA and protein when compared to nuclear translocation of ER alone. Previous reports have demonstrated that a combination of a promoter sequence and an enhancer region located in the first intron of Csf1r gene (Fms intronic regulatory element: FIRE) is enough to recapitulate the endogenous Csf1r expression and that these elements contained consensus binding sites for C/EBP transcription factors. Then, we hypothesized that C/EBPβ binds to these sites, activates transcription of Csf1r gene and promotes survival of Ly6C- monocytes. To evaluate this hypothesis, we utilized an expression vector, in which green fluorescent protein (GFP) is driven by a combination of the Csf1r promoter and FIRE sequences (Csf1r-EGFP-FIRE) (a kind gift from Drs Clare P and David A Hume, University of Edinburgh). When a C/EBPβexpression vector was co-transfected with the vector containing Csf1r-EGFP-FIRE into HEK293 cells, the frequencies of GFP positive cells were significantly higher when compared to a control vector (C/EBPβ vs control; 4.6±0.6 vs 1.6±1.0, p=0.01), suggesting that C/EBPβ regulates Csf1r expression through these elements. We are currently evaluating the significance of C/EBP consensus binding sites in the promoter and the enhancer. ChIP PCR is also in progress to further verify our hypothesis. Collectively, these results suggest that Csf1r is a critical downstream target of C/EBPβ in Ly6C- monocytes. Disclosures No relevant conflicts of interest to declare.

scholarly journals Evolutionarily conserved transcriptional landscape of the heart defining the chamber specific physiology

2021 ◽  
Author(s):  
Shrey Gandhi ◽  
Anika Witten ◽  
Federica deMajo ◽  
Martijn Gilbers ◽  
Jos Maessen ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Expression Patterns ◽  
Circular Rna ◽  
Model Organisms ◽  
Protein Coding ◽  
Evolutionarily Conserved ◽  
Cardiovascular Biology ◽  
Novel Therapeutic Strategies ◽  
Transcriptional Landscape

AbstractCardiovascular disease (CVD) remains the leading cause of death worldwide. A deeper characterization of the regional transcription patterns within different heart chambers may aid to improve our understanding of the molecular mechanisms involved in the function of the heart as well as our ability to develop novel therapeutic strategies. Here, we determined differentially expressed protein coding, long non-coding (lncRNA) and circular RNA (CircRNA) genes within various heart chambers across seven vertebrate species. We identified chamber specific genes, lncRNAs and pathways that are evolutionarily conserved in vertebrates. Further, we identified lncRNA homologs based on sequence, secondary structure, synteny and expressional conservation. Interestingly, most lncRNAs were found to be syntenically conserved. Various factors affect the co-expression patterns of transcripts including (i) genomic overlap, (ii) strandedness and (iii) transcript biotype. We also provide a catalogue of CircRNAs which are abundantly expressed across vertebrate hearts. Finally, we established a repository called EvoACTG (http://evoactg.uni-muenster.de/), which provides information about the conserved expression patterns for both PC genes and non-coding RNAs (ncRNAs) in the various heart chambers, and may serve as a community resource for investigators interested in the (patho)-physiology of CVD. We believe that this study will inform researchers working in the field of cardiovascular biology to explore the conserved yet intertwined nature of both coding and non-coding cardiac transcriptome across various popular model organisms in CVD research.

scholarly journals Single Cell Transcriptome-Based Dissection of Lineage Fate Decisions in Myelopoiesis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1395-1395
Author(s):  
Andre Olsson ◽  
H. Leighton Grimes ◽  
Virendra K Chaudhri ◽  
Philip Dexheimer ◽  
Bruce J Aronow ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Fate ◽  
Molecular Mechanisms ◽  
Conflicts Of Interest ◽  
Expression Patterns ◽  
Rna Seq ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome ◽  
Cell Data ◽  
Insight Into

Abstract In spite of tremendous advances in the analysis of hematopoietic progenitors and transcription factors that give rise to different lineages, molecular insight into the mechanisms that underlie cell fate choice at the level of individual cells is lacking. We utilized single-cell RNA sequencing of murine granulocyte-monocyte progenitors (GMPs) to analyze the molecular basis of cell fate choice. Over 200 libraries were generated with average read depths of 4 million per library and an expressed gene call of over 3,800 genes with FPKM >3. Our data reveal a varied but coherent spectrum of gene expression patterns in individual murine GMPs. The majority of cells could be clustered into ones expressing either granulocytic or monocytic genes, suggesting that they were primed for lineage determination. A minority of GMPs expressed a mixed-lineage pattern of genes. The single-cell data suggested an antagonistic transcription factor circuit involving Gfi1 and IRF8 that was validated with both loss- and gain-of-function experiments in GMPs. Our data highlight the utility of single cell RNA-Seq analysis to reveal molecular mechanisms controlling lineage fate decisions in hematopoiesis. Disclosures No relevant conflicts of interest to declare.

Establishment and Characterization of A New Leukemia Mouse Model Induced by MLL/AF4 Fusion Protein.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3573-3573
Author(s):  
Hayato Tamai ◽  
Koichi Miyake ◽  
Noriko Miyake ◽  
Hiroki Yamaguchi ◽  
Kazuo Dan ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Molecular Mechanisms ◽  
Conflicts Of Interest ◽  
Fusion Gene ◽  
Lymphoblastic Leukemia ◽  
Mixed Lineage Leukemia ◽  
Lymphoid Cells ◽  
Long Latency ◽  
Infant Leukemia

Abstract Abstract 3573 Poster Board III-510 Background MLL/AF4 fusion is the most common abnormality in infant leukemia and is associated with a poor prognosis. Although further study and treatment should be groped for acute lymphoblastic leukemia (ALL) with MLL/AF4, the lack of a proper animal model has impeded understanding of the molecular mechanism of leukemia associated with the MLL/AF4 fusion gene. Previous mouse models suggest that the tumorigenesis capacity of MLL/AF4 alone is insufficient for development mixed-lineage leukemia. MLL/AF4 is capable of inducing lymphoid malignancy with long latency, which is very different from aggressive pro-B ALL in humans. Recently, we have reported that enhancement of S100A6 (calcyclin) expression played an important role in MLL/AF4-leukemogenesis using murine 32Dc cell line. In this study, we tried to make improved MLL/AF4 transgenic (Tg) mice which reflect human leukemogenicity and analyzed molecular mechanism of MLL/AF4 fusion gene. Methods To get high and sustained expression of MLL/AF4 in hematological lineage, we used MSCV promoter to express MLL/AF4 fusion gene. After MLL/AF4 expression plasmids were injected into fertilized eggs of mice (C57BL/6), the manipulated embryos were transferred into the oviducts of pseudopregnant females to make MLL/AF4 Tg mice. Finally we characterized MLL/AF4 Tg mice and analyzed molecular mechanism of MLL/AF4-leukemogenesis. Results Our new established MLL/AF4 Tg mice had lymphoma earlier than previous MLL/AF4 knock-in mice (median 12M vs 17M). Infiltration of lymphoid cells was located in liver, lung, kidney, and spleen and they consist of pro B-cell (CD45R+CD43+CD19-) lineage. Leukemic change developed two months (median 14M) after appearance of lymphoma. More than 30% of abnormal lymphocytes and severe anemia (RBC: 276×104 ml, Hb: 4.9 g/ml) were found in peripheral blood in MLL/AF4 Tg mice. Western blot analysis showed that up-regulation of HoxA9 and S100A6 was detected in MLL/AF4 Tg mouse compared to wild type C57BL/6. Conclusion We succeeded in establishment of improved more aggressive MLL/AF4 Tg mice which reflect MLL/AF4 leukemogenicity of humans. Appearance of lymphoma in our MLL/AF4 Tg mice needed short er latency than those of the previous knock-in mice. Enhancement of HoxA9 and S100A6 expression was involved in MLL/AF4-associated leukemogenesis. We believe that this new MLL/AF4 Tg mouse is useful for study of the molecular mechanisms of MLL/AF4 leukemogenesis and development of new therapy for mixed-lineage leukemia. Disclosures: No relevant conflicts of interest to declare.

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