Leukemia Transcription Factor Research Articles

PBX1: A Novel Stage-Specific Regulator of Adipocyte Development

Although adipocyte terminal differentiation has been extensively studied, the early steps of adipocyte development and the embryonic origin of this lineage remain largely unknown. Here we describe a novel role for the pre-B-cell leukemia transcription factor one (PBX1) in adipocyte development using both mouse embryonic stem cells (mESCs) and human multipotent adipose-derived stem (hMADS) cells. We show that Pbx1(-/-) mESCs are unable to generate adipocytes, despite normal expression of neuroectoderm and neural crest (NC) markers. Early adipocyte lineage markers are not induced in Pbx1(-/-) mESCs, suggesting that Pbx1 controls the generation and/or the maintenance of adipocyte progenitors (APs) from the NC. We further characterize the function of PBX1 in postnatal adipogenesis and show that silencing of PBX1 expression in hMADS cells reduces their proliferation by preventing their entry in the S phase of the cell cycle. Furthermore, it promotes differentiation of hMADS cells into adipocytes and partially substitutes for glucocorticoids and rosiglitazone, two key proadipogenic agents. These effects involve direct modulation of PPARγ activity, most likely through regulation of the biosynthesis of PPARγ natural endogenous ligand(s). Together, our data suggest that PBX1 regulates adipocyte development at multiple levels, promoting the generation of NC-derived APs during embryogenesis, while favoring APs proliferation and preventing their commitment to the adipocyte lineage in postnatal life.

Deregulated transcription factors in leukemia

Specific chromosomal translocations and other mutations associated with acute myeloblastic leukemia (AML) often involve transcription factors and transcriptional coactivators. Such target genes include AML1, C/EBPα, RARα, MOZ, p300/CBP, and MLL, all of which are important in the regulation of hematopoiesis. The resultant fusion or mutant proteins deregulate the transcription of the affected genes and disrupt their essential role in hematopoiesis, causing differentiation block and abnormal proliferation and/or survival. This review focuses on such transcription factors and coactivators, and describes their roles in leukemogenesis and hematopoiesis.

Role of Meis1 in mitochondrial gene transcription of pancreatic cancer cells

Pre-B-cell leukemia transcription factor (PBX)/three-amino-acid loop extension (TALE) class transcription factors [PBX1–4, Meis homeobox (Meis) 1–3, pbx/knotted 1 homeobox (Prep) 1, 2] are involved in tumorigenesis and metastasis. To investigate further the function of PBX/TALE class transcription factors, mRNA expression profile after downregulation of each mRNA expression by siRNA transfection in pancreatic cancer cell line, Panc-1, was examined. Downregulation of Meis1 resulted in downregulation of mitochondrial genes, but those of PBX1 and PBX2 did not. Quantitative reverse transcription polymerase chain reaction confirmed downregulation of mitochondrial genes by Meis1 siRNA transfection. Chromatin immunoprecipitation assay revealed the binding of Meis1 to the mitochondrial promoter region that contained the putative Meis1 binding site. Luciferase reporter assay showed the increase of luciferase activity of a construct containing the Meis1 binding site compared with that with shorter fragment without Meis1 binding region. These findings indicate that Meis1 works as a transcription factor for mitochondrial genes in pancreatic cancer cells.

SMADs and FOXL2 Synergistically Regulate Murine FSHβ Transcription Via a Conserved Proximal Promoter Element

Pituitary FSH regulates ovarian and testicular function. Activins stimulate FSHβ subunit (Fshb) gene transcription in gonadotrope cells, the rate-limiting step in mature FSH synthesis. Activin A-induced murine Fshb gene transcription in immortalized gonadotropes is dependent on homolog of Drosophila mothers against decapentaplegic (SMAD) proteins as well as the forkhead transcription factor FOXL2 (FOXL2). Here, we demonstrate that FOXL2 synergizes with SMAD2, SMAD3, and SMAD4 to stimulate murine Fshb promoter-reporter activity in heterologous cells. Moreover, SMAD3-induction of Fshb promoter activity or endogenous mRNA expression is dependent upon endogenous FOXL2 in homologous cells. FOXL2/SMAD synergy requires binding of both FOXL2 and SMAD3 or SMAD4 to DNA. Of three putative forkhead-binding elements identified in the murine Fshb promoter, only the most proximal is absolutely required for activin A induction of reporter activity in homologous cells. Additionally, mutations to the minimal SMAD-binding element adjacent to the proximal forkhead-binding element abrogate activin A or FOXL2/SMAD3 induction of reporter activity. In contrast, a mutation that impairs an adjacent PBX1/PREP1 (pre-B cell leukemia transcription factor 1-PBX/knotted-1 homeobox-1) binding site does not alter activin A-stimulated promoter activity in homologous cells. Collectively, these and previous data suggest a model in which activins stimulate formation of FOXL2-SMAD2/3/4 complexes, which bind to the proximal murine Fshb promoter to stimulate its transcription. Within these complexes, FOXL2 and SMAD3 or SMAD4 bind to adjacent cis-elements, with SMAD3 brokering the physical interaction with FOXL2. Because this composite response element is highly conserved, this suggests a general mechanism whereby activins may regulate and/or modulate Fshb transcription in mammals.

Coupled Folding and Binding of pKID with KIX Domain Investigated by Multicanonical Molecular Dynamics Simulation in Explicit Solvent

Intrinsically disordered proteins (IDPs) do not form any rigid tertiary structures alone. Most of them bind corresponding proteins and fold into an ordered structure to play important roles in biological functions such as signal transduction cascades. The phosphorylated kinase induced domain (pKID) is one of IDPs. The pKID adapts an ordered helical structure while binds to the KIX domain. This structural property of pKID is called “coupled folding and binding”. From nuclear magnetic resonance (NMR) studies a kinetics model of the coupled folding and binding has been proposed: there are four states of pKID (disordered, encounter complex, intermediate, and ordered helical structure). However, the details of these states at atomic level were still unclear.In order to obtain the free energy landscapes and the stable complex structures at various temperatures for the system, where pKID, the KIX domain, water and ions are included and they can interact with each other, multicanonical molecular dynamics (McMD) simulation was performed with the all-atom model in explicit solvent. McMD simulation is one of generalized ensemble methods, which can search conformational space much wider than a conventional molecular dynamics simulation, as well as the replica-exchange molecular dynamics (REMD). In this study, starting from completely disordered and unbound states of pKID, the helices of pKID were reproduced as approaching to the KIX domain at adequate low temperature. Additionally, we found a different binding mode from the NMR model. Interestingly, this binding mode is similar to that of the activation domain of the mixed lineage leukemia (MLL) transcription factor upon binding to the KIX domain.

Prognostic significance of pre B cell leukemia transcription factor 2 (PBX2) expression in non‐small cell lung carcinoma

Previous studies on the mammary carcinoma cell line have shown that the pre B cell leukemia transcription factor 1 (PBX1) was a transcription factor for valosin-containing protein (VCP), which is involved in invasion and metastasis of cancers. The roles of PBX1 and PBX2, a highly homologous transcription factor to PBX1, for expression of VCP were examined in the cell lines from non-small cell lung cancer (NSCLC). The effects of PBX1 and PBX2 on VCP expression were examined with siRNA in A549 and PC14 NSCLC cell lines. Expression levels of PBX2 and VCP were immunohistochemically examined and compared with each other in 206 NSCLC cases. Subsequently, significance of PBX expression in clinical behavior of NSCLC patients was evaluated. Expression levels of VCP mRNA significantly decreased when PBX2 but not PBX1 expression was knocked down in NSCLC cell lines. Immunohistochemically, staining intensity of PBX2 was correlated with that of VCP in clinical samples. Then correlation of PBX2 expression and clinical behavior of NSCLC patients was evaluated. Univariate analysis revealed high expression levels of PBX2 and VCP to be poor prognosticators for overall and disease-free survival. Multivariate analysis revealed that high expression of VCP but not PBX2 to be an independent prognostic factor. PBX2 is a transcription factor for VCP in NSCLC. Because high levels of PBX2 expression correlated with prognosis of NSCLC, PBX2 could be a target molecule for treatment of NSCLC.

Ikaros family transcription factors in chronic and acute leukemia

lkaros, Aiolos, and Helios are transcription factors important in lymphocyte differentiation and development [1-6]. One of the most interesting aspects of the Ikaros family members is that they are expressed as multiple splice variants and proteins arising from alternatively spliced transcripts may act as dominant negatives. Although expression of these short isoforms has been linked to lymphoid and myeloid leukemia [7-11] the comparison of Ikaros members mRNA expression level has not been reported. Here we show, by means of real time qRT-PCR method, a quantitative distribution of Ikaros, Aiolos, and Helios mRNA in hemopoietic cells of patients with lymphocytic leukemia, represented by the most frequent B cell chronic lymphocytic leukemia (CLL) and acute lymphocytic leukemia (T cell ALL and common ALL), and compared with the acute leukemia of the myeloid lineage, acute myeloid leukemia (AML). We found that the relative quantity of Ikaros and Helios mRNA was similar in all subgroups. However, analysis of Aiolos showed a clear difference among groups because of its lower expression in all types of acute leukemia investigated. Our results associating a lower Aiolos expression with acute leukemia point to Aiolos's role in human lymphocyte development, as previously shown in mice.

Direct Observation of the Dynamic Process Underlying Allosteric Signal Transmission

Allosteric regulation is an effective mechanism of control in biological processes. In allosteric proteins a signal originating at one site in the molecule is communicated through the protein structure to trigger a specific response at a remote site. Using NMR relaxation dispersion techniques we directly observe the dynamic process through which the KIX domain of CREB binding protein communicates allosteric information between binding sites. KIX mediates cooperativity between pairs of transcription factors through binding to two distinct interaction surfaces in an allosteric manner. We show that binding the activation domain of the mixed lineage leukemia (MLL) transcription factor to KIX induces a redistribution of the relative populations of KIX conformations toward a high-energy state in which the allosterically activated second binding site is already preformed, consistent with the Monod-Wyman-Changeux (WMC) model of allostery. The structural rearrangement process that links the two conformers and by which allosteric information is communicated occurs with a time constant of 3 ms at 27 degrees C. Our dynamic NMR data reveal that an evolutionarily conserved network of hydrophobic amino acids constitutes the pathway through which information is transmitted.

The pathogenetic significance of deregulated hematopoietic transcription factors in leukemia

The pathogenetic significance of deregulated hematopoietic transcription factors in leukemia

Gene Alterations and Expression of Hematopoietic Transcription Factors in Pediatric Acute Megakaryoblastic Leukemia (AMKL): A Comparison of Down Syndrome (DS)-Associated AMKL with Non-DS-AMKL.

Morphological and cytogenetic studies have demonstrated there is a diverse heterogeneity for acute megakaryoblastic leukemia (AMKL). In children, there are 2 major AMKL subgroups, with the disease developing in patients with and without Down syndrome (DS-AMKL and non-DS-AMKL, respectively). Somatic mutations of the GATA1 gene are found in children with DS-AMKL. Recently, activating mutations of the JAK3 gene have been found in both DS-AMKL and non-DS-AMKL patients. Furthermore, JAK2V617F plays a critical role in the pathogenesis of myeloproliferative disorders that are found in adults with non-DS-AMKL. To examine the biological characteristics of the DS-AMKL and non-DS-AMKL blast cells, we compared the morphology, immunophenotypes, gene alterations (GATA1, JAK2, JAK3) and the expression of the hematopoietic transcription factors in blast cells derived from 29 DS-AMKL and 21 non-DS-AMKL children. AMKL was defined by the presence of blast cells that expressed at least one platelet-associated antigen (CD36, CD41, CD42, or CD61). Written informed consent for banking and molecular analysis of the blast cells was obtained from the parents of each of the patients. Blast cell morphology was categorized as follows: type 1, completely undifferentiated blasts; type 2, intermediately differentiated blasts with cytoplasmic blebs; type 3, blasts with dysmegakaryocytopoiesis; and type 4, blasts with deep blue cytoplasm. The respective distributions of the morphology of the blasts in DS-AMKL and non-DS-AMKL patients were as follows: type 1 (41%, 39%), type 2 (4%, 39%), type 3 (11%, 22%), and type 4 (44%, 0%). Type 4 blasts were only seen in DS-AMKL. Atypical expression of lymphoid-associated antigen CD7 was detected in 90% and 53% of the immunophenotyped-patients with DS-AMKL and non-DS-AMKL, respectively (p = 0.003). Glycophorin A was only detected on the blasts of 34% of the patients with DS-AMKL (p = 0.012). Interestingly, 78% of the type 4 blasts were positive for glycophorin A. The GATA1, JAK2 and JAK3 mutations were analyzed in bone marrow or peripheral blood samples. High-molecular weight DNA was extracted from the samples using standard methods. We amplified the genomic DNA corresponding to exon2 of GATA1, exon14 of JAK2, and all 23 exons of the JAK3 gene, respectively, with the amplified products sequenced directly on a DNA sequencer (310; Applied Biosystems, Foster City, CA) using a BigDye terminator cycle sequencing kit (Applied Biosystems). The frequencies of the gene mutations in DS-AMKL and non-DS-AMKL were distributed as follows: GATA1 (100%, 8%), JAK2V617F (6%, 0%) and JAK3 (12%, 8%). To define the differentiation stage based on the molecular level, we quantified the expression of the transcription factors related to the megakaryo-erythroid cell lineages. The expression of the transcription factors (GATA1, GATA2, FOG-1, NF-E2, SCL, PU.1, C/EBPa, c-mpl, Epo-R, CD41b, bglobin) was analyzed by real time RT-PCR in blasts that were sorted by flow cytometry. Total RNA was extracted from the purified blasts. Synthesis of cDNA was performed using a Thermoscript RT-PCR system (Invitrogen, San Diego, CA) according to the manufacturer's instructions. Real-time RT-PCR was performed using an ABI Prism 7000 Sequence Detection System (Applied Biosystems, Branchburg, NJ). Normalized gene expression levels are given as the ratio between the mean value for the target gene and that for the GAPDH gene in each sample. Median bglobin transcript levels in DS-AMKL blasts were significantly higher than those seen for the non-DS-AMKL blasts (p = 0.006). Particularly, median bglobin and GATA1 transcript levels in the type 4 blasts, which were only found in DS-AMKL, were significantly higher than those found in the blasts of other types (p = 0.033 and p = 0.021, respectively). Median GATA1, NF-E2 and Epo-R transcript levels in type 3 blasts, which differentiated into mature megakaryocytes, were significantly lower than those found in the blasts of other types (p = 0.030, p = 0.036, and p = 0.028, respectively). In conclusion, blasts of DS-AMKL differentiated not only into the megakaryocyte but also the erythroid lineage because of the overexpression of glycophorin A and bglobin under the GATA1 gene mutations. Considering the totally different gene alteration patterns and the levels of transcription factors that were observed, DS-AMKL needs to be classified into a category that differs from non-DS-AMKL.

CBP/p300-Interacting Transactivator, with Glu/Asp-Rich C-Terminal Domain, 2, and Pre-B-Cell Leukemia Transcription Factor 1 in Human Adrenal Development and Disease

Disorders of adrenal development result in significant morbidity and mortality. However, the molecular basis of human adrenal development, and many forms of disease, is still poorly understood. We evaluated the role of two new candidate genes, CBP/p300-interacting transactivator, with Glu/Asp-rich C-terminal domain, 2 (CITED2), and pre-B-cell leukemia transcription factor 1 (PBX1), in human adrenal development and disease. CITED2 and PBX1 expression in early human fetal adrenal development was assessed using RT-PCR and in situ hybridization. The regulation of CITED2 and PBX1 by steroidogenic factor-1 (SF-1) and dosage-sensitive sex reversal, adrenal hypoplasia congenital, critical region on the X chromosome, gene-1 (DAX1) was evaluated in NCI-H295R human adrenocortical tumor cells by studying promoter regulation. Finally, mutational analysis of CITED2 and PBX1 was performed in patients with primary adrenal disorders. CITED2 and PBX1 are expressed in the human fetal adrenal gland during early development. Both genes are activated by SF-1 in a dose-dependent manner in NCI-H295R cells, and, surprisingly, PBX1 is synergistically activated by SF-1 and DAX1. Mutational analysis failed to reveal significant coding sequence changes in individuals with primary adrenal disorders. CITED2 and PBX1 are likely to be important mediators of adrenal development and function in humans, but mutations in these genes are not common causes of adrenal failure in patients in whom a molecular diagnosis remains unknown. The positive interaction between DAX1 and SF-1 in regulating PBX1 may be an important mechanism in this process.

Expression of pre B cell leukemia transcription factor (PBX) is correlated with prognosis and survival of patients with non-small cell lung carcinoma

22222 Background: Pre B cell leukemia transcription factor 1 (PBX1) is a transcription factor for valosin-containing protein (VCP), which is involved in invasion and metastasis of cancers. To examine the prognostic significance of PBX1 and other PBX members, PBX2 and PBX3, in non-small cell lung cancer (NSCLC), the present study was designed. Methods: Expression of PBX1, PBX2, and PBX3 was immunohistochemically examined in 206 NSCLC patients (132 men and 74 women) with ages ranging from 35 to 78 (median, 62). Staining intensity of tumor cells was categorized as equal to or stronger expression than endothelial cells in less than 20% (score 1) or more than 20% (score 2). Results: PBX2 but not PBX1 and PBX3 nuclear expression correlated with high expression of VCP. Univariate analysis revealed score 1 PBX1 cytoplasmic expression, score 2 PBX2 nuclear expression, score 2 PBX2 cytoplasmic expression to be poor prognosticators for overall and disease-free survival. Multivariate analysis revealed that PBX1 cytoplasmic expression, visceral pleural invasion, and tumor size as independent prognostic factors for disease-free survival, and PBX1 cytoplasmic expression, PBX2 cytoplasmic expression, VCP expression, age, and visceral pleural invasion for overall survival. Conclusions: The present study showed that increased expression of PBX2 and decreased expression of PBX1 were poor prognosticators for NSCLC. Close correlation between PBX2 nuclear expression and VCP expression suggested the role of PBX2 as a transcription factor controlling VCP expression. No significant financial relationships to disclose.

The association of PBX1 polymorphisms with overweight/obesity and metabolic alterations in the Korean population

Pre-B-cell leukemia transcription factor 1 (PBX1), which is located on chromosome 1q23, was recently reported to be associated with type 2 diabetes mellitus. We examined whether single nucleotide polymorphisms (SNPs) of the PBX1 gene are associated with overweight/obesity in a Korean population. We genotyped 66 SNPs in the PBX1 gene and investigated their association with clinical phenotypes found in 214 overweight/obese subjects and 160 control subjects using the Affymetrix Targeted Genotyping chip array. Seven SNPs (g.+75186C>T, g.+78350C>A, g.+80646C>T, g.+138004C>T, g.+185219G>A, g.+191272A>C, and g.+265317T>A) were associated with the risk of obesity in three models (codominant, dominant, and recessive) (P=0.007-0.05). Haplotype 1 (CAC) and 3 (TAC) of block 3 and haplotype 2 (GGAAT) of block 10 were also strongly associated with the risk of obesity. In the control group, subjects that had homozygote for the major allele for both g.+185219G>A and g.+191272A>C showed lower high density lipoprotein-cholesterol (HDL-C) level compared to those possessing the minor allele, suggesting that the association between the homozygote for the major allele for both g.+185219G>A and g.+191272A>C and HDL-C is attributable to the increased risk of obesity. This study suggests that the PBX1 gene is a possible risk factor in overweight/obese patients.

Vitamin E Isoforms Inhibit Cell Proliferation and Downregulate Homeobox Protein Expression in the Leukemic KG-1 Cells.

Vitamin E is a fat-soluble vitamin that exists in eight different isoforms: four tocopherols (alpha, beta-, gamma- and delta-), and four tocotrienols (also alpha-, beta-, gamma- and delta-). Studies are underway to determine whether vitamin E, through its ability to limit production of free radicals, might help prevent or delay the development of cancer. Other studies suggest that vitamin E may modulate signal transduction pathways to prevent or reverse the effects of cancer. In our study, we examined the effect of vitamin E isotypes on some gene products involved in acute leukemia such as HOXA9 (Homeobox A9), PBX1 (Pre-B cell leukemia transcription factor 1) and E2A-PBX1. HOXA9 is a transcription factor with a central role in both hemopoiesis and leukemia. A high level of HOXA9 is a characteristic feature of acute myeloid leukemia (AML) and may be sufficient to cause this disease. PBX1 is involved in a form of pre-B-cell acute lymphoblastic leukemia (B-ALL). E2A-PBX1 is a protein resulting from a chromosomal translocation t(1;19)(q23;p13.3) which involves PBX1 and E2A genes. E2A-PBX1 transforms cells by constitutively activating transcription of genes regulated by PBX1 or by other members of the PBX protein family. KG-1 cells were treated with 20 micromolar (μM) gamma tocotrienol (γT3) and delta tocotrienol (δT3) for 48 hours, and 50 μM gamma tocopherol (γT) and delta tocopherol (δT) for 72 hours. MTT Assays demonstrate 30% inhibition of cell proliferation using 50 μM γT after 72 hours, 60% inhibition of cell proliferation using 50 μM δT after 48 hours, 70% inhibition of cell proliferation using 25 μM γT3 after 48 hours, and 75% inhibition of cell proliferation using 25 μM δT3 after 48 hours. Western Blot analysis was performed on the cell lysates using HOXA9, PBX1 and E2A-PBX-1 antibodies. Our results showed that both δT and γT when used at a 50 μM concentration for 72 hours treatment, as well as δT3 and γT3 when used at a 20 μM concentration for 48 hours treatment, have a down regulatory effect on HOXA9, PBX1 and E2A-PBX-1 protein expression in the leukemic KG-1 cells. A down regulatory effect on cyclin D1 and cyclin D3 was also demonstrated.

Targeting Transcription Factors in Acute Leukemia in Children

Transcription factors play essential roles in controlling normal blood development and their alteration leads to abnormalities in cell proliferation, differentiation and survival. In many childhood acute leukemias, transcription factors are altered through chromosomal translocations that change their functional properties resulting in repressed activity or inappropriate activation. The development of therapies that specifically target these molecular abnormalities holds promise for improving the outcome in diseases that remain challenging to treat, such as childhood T-cell acute lymphoblastic leukemia and acute myeloid leukemia, with improved toxicity profiles. All trans-retinoic acid and arsenic trioxide have already demonstrated efficacy in acute promyelocytic leukemia in both adults and children. Newer agents, such as histone deacetylase inhibitors, drugs targeting the NOTCH pathway, and short interfering RNAs have shown encouraging results in pre-clinical studies and are likely to enter the clinical arena in the near future. Through an improved understanding of the pathways and mechanisms underlying the malignant transformation induced by altered transcription factors, new targeted therapies will be designed that should greatly enhance current available treatments.

In the Forefront of Basic and Translational Cancer Research: Seventh Joint Conference of the American Association for Cancer Research and the Japanese Cancer Association

The Seventh Joint Conference of the American Association of Cancer Research and the Japanese Cancer Association, “In the Forefront of Basic and Translational Cancer Research,” held in Waikoloa, HI, on January 21 to 25, 2007, was designed to showcase the latest advances in all major areas of

Pre-B-Cell Leukemia Transcription Factor 1 Regulates Expression of Valosin-Containing Protein, a Gene Involved in Cancer Growth

Valosin-containing protein (VCP) is involved in a wide variety of cellular functions. Our previous studies showed that the enhanced expression of VCP in cancer cells correlated with invasion and metastasis of cancers. Here, the regulatory mechanism for VCP transcription was investigated. Luciferase reporter constructs containing serially deleted 5'-flanking region of the VCP gene were transfected into MCF7 mammary carcinoma cell line, in which VCP was abundantly expressed. The deletion and mutation at the two binding motifs for pre-B-cell leukemia transcription factor 1 (PBX1) reduced the luciferase activity, indicating that these two PBX1 motifs mediated the transactivation of the VCP gene. Chromatin immunoprecipitation assay showed the binding of PBX-1 to the 5'-flanking region of the VCP gene. The knockdown of PBX1 by siRNA decreased the expression level of VCP. VCP is reported to maintain cell viability after the treatment of tumor necrosis factor-alpha. The viability of tumor necrosis factor-alpha-treated cells was significantly reduced in PBX1 knockdown MCF7. These findings indicate that PBX1 plays a crucial role in VCP expression and function and that the PBX-VCP pathway might be important for cell survival under cytokine stress.

The pathogenetic significance of deregulated hematopoietic transcription factors in leukemia

The pathogenetic significance of deregulated hematopoietic transcription factors in leukemia

The PDX1 Homeodomain Transcription Factor Negatively Regulates the Pancreatic Ductal Cell-specific Keratin 19 Promoter

Keratin 19 is a member of the cytokeratin family that is critical for maintenance of cellular architecture and organization, especially of epithelia. The pancreas has three distinct cell types, ductal, acinar, and islet, each with different functions. Embryologically, the pancreatic and duodenal homeobox 1 (PDX1) homeodomain protein is critical for the initiation of all pancreatic lineages; however, the later differentiation of the endocrine pancreas is uniquely dependent upon high PDX1 expression, whereas PDX1 is down-regulated in the ductal and acinar cell lineages. We find that this down-regulation may be required for normal ductal expression of cytokeratin K19. The K19 promoter-reporter gene assay demonstrates that ectopic PDX1 inhibits K19 reporter gene activity in primary pancreatic ductal cells. This is reinforced by our findings that retrovirally mediated stable transduction of PDX1 in primary pancreatic ductal cells suppresses K19 expression, and short interfering RNA to PDX1 in Min6 insulinoma cells results in the induction of normally undetectable K19. Complementary functional and biochemical approaches led to the unexpected finding that a multimeric complex of PDX1 and two members of the TALE homeodomain factor family, MEIS1a and PBX1b, regulates K19 gene transcription through a specific cis-regulatory element (-341 to -325) upstream of the K19 transcription start site. These data suggest a unifying mechanism whereby PDX1, myeloid ecotropic viral insertion site (MEIS), and pre-B-cell leukemia transcription factor 1 (PBX) may regulate ductal and acinar lineage specification during pancreatic development. Specifically, concomitant PDX1 suppression and MEIS isoform expression result in proper ductal and acinar lineage specification. Furthermore, PDX1 may inhibit the ductal differentiation program in the pancreatic endocrine compartment, particularly beta cells.

Association evidence of schizophrenia with distal genomic region of NOTCH4 in Taiwanese families

Evidence for association with schizophrenia has been reported for NOTCH4, although results have been inconsistent. Previous studies have focused on polymorphisms in the 5' promoter region and first exon of NOTCH4. Our aim was to test the association of the entire genomic region of NOTCH4 in 218 families with at least two siblings affected by schizophrenia in Taiwan. We genotyped seven single nucleotide polymorphisms (SNPs) of this gene, with average intermarker distances of 5.3 kb. Intermarker linkage disequilibrium (LD) was calculated using gold software, and single-locus and haplotype association analyses were performed using transmit software. We found that the T allele of SNP rs2071285 (P= 0.035) and the G allele of SNP rs204993 (P= 0.0097) were significantly preferentially transmitted to the affected individuals in the single-locus association analysis. The two SNPs were in high LD (D' > 0.8). Trend for overtransmission was shown for the T-G haplotype of the two SNPs to affected individuals (P= 0.053), with the A-A haplotype significantly undertransmitted (P= 0.034). The associated region distributed across the distal portion of the NOTCH4 gene and overlapped with the genomic region of the G-protein signaling modulator 3 and pre-B-cell leukemia transcription factor 2. In summary, we found modest association evidence between schizophrenia and the distal genomic region of NOTCH4 in this Taiwanese family sample. Further replication for association with the distal genomic region of NOTCH4 is warranted.