Mutations In Transcription Factor Genes Research Articles

Clonal Evolution Pattern and Prognostic Significance of Clonal Architecture in KMT2A-Rearranged Acute Myeloid Leukemia

MLL (KMT2A) rearrangements are among the most frequent chromosomal abnormalities that occur in acute myeloid leukemia (AML). Mutational landscapes in KMT2A-rearranged AML have been reported; however, most studies are missing data at relapse. Therefore, matched diagnostic and relapse samples were analyzed in this study, and the clonal evolution pattern in KMT2A-rearranged AML was examined. Further, the prognostic significance of the clonal architecture was investigated.Sixty-two diagnostic and 16 relapse samples obtained from pediatric patients with KMT2A-rearranged AML enrolled in the Japan Children's Cancer Group (JCCG) AML-05/AML-99 study were analyzed for 338 genes using targeted sequencing. The data were analyzed with the published data of 105 diagnostic and 9 relapse samples with KMT2A-rearranged AML. Additionally, as a control, the mutation data of matched diagnostic and relapse samples of 107 patients with non-KMT2A-rearranged AML were collected.Among 25 patients with KMT2A-rearranged AML with matched data at diagnosis and relapse, mutations of signaling pathway genes (FLT3, KRAS, NRAS, PTPN11, CBL, and BRAF) were frequently detected in diagnostic samples (25 mutations/25 patients). However, 21 of 25 (84.0%) mutations were lost at relapse. In contrast, 7 of 19 (36.8%) mutations of other pathway genes were lost at relapse, and the percentage was significantly lower than that of mutations in the signaling pathway genes (P = 0.002). Six mutations in the signaling pathway genes and 11 mutations in other pathway genes were acquired at relapse. Particularly, mutations of transcription factor genes (WT1, SPI1, GATA2, and RUNX1) were acquired at relapse: 7 of 8 (87.5%) mutations were detected only at relapse. These results suggest that mutations of signaling pathway genes are unstable in the clonal evolution of KMT2A-rearranged AML. Mutations of other pathway genes, especially those of transcription factor genes, may contribute to relapse in patients with KMT2A-rearranged AML.Next, attention was turned to the KRAS mutations (KRAS-MT) because we have previously shown that KRAS-MT are independent adverse prognostic factors in KMT2A-rearranged AML (Blood Adv. 2020). Among 25 patients with KMT2A-rearranged AML with matched data at diagnosis and relapse, 10 (40.0%) patients harbored KRAS-MT at diagnosis. Interestingly, KRAS-MT were lost at relapse in 9 of 10 (90.0%) patients. Among 107 patients with non-KMT2A-rearranged AML with matched data at diagnosis and relapse, 10 (9.3%) patients harbored KRAS-MT at diagnosis. The frequency of KRAS-MT was significantly higher in KMT2A-rearranged AML (40.0% vs. 9.3%, P = 0.0006). This may be explained on the basis of the fact that KRAS-MT is associated with a high relapse rate in KMT2A-rearranged AML, but not in non-KMT2A-rearranged AML. KRAS-MT was lost at relapse in 5 of 10 (50.0%) patients with non-KMT2A-rearranged AML. The percentage of KRAS-MT loss at relapse was higher in KMT2A-rearranged AML. However, it was not statistically significant (90.0% vs. 50.0%, P = 0.14). Therefore, KRAS-MT may be unstable in clonal evolution regardless of disease subtypes in AML. The underlying mechanisms of the paradox between the high relapse rate in patients with KRAS-MT and frequent loss of KRAS-MT at relapse in patients with KMT2A-rearranged AML should be examined in future studies.The loss of KRAS-MT at relapse suggests that the mutations were in subclones at diagnosis. Therefore, we finally examined the prognosis of 167 patients according to the clonality of KRAS-MT at diagnosis. In patients with KMT2A-MLLT3 (n = 67), those with subclonal KRAS-MT (n = 6) had adverse 5-y event-free survival compared with both patients with wild-type KRAS (KRAS-WT) (n = 56) (KRAS-WT vs. subclonal KRAS-MT: 58.7% vs. 16.7%, P = 0.04) and patients with clonal KRAS-MT (n = 5) (clonal KRAS-MT vs. subclonal KRAS-MT: 80.0% vs. 16.7%, P = 0.07). However, 5-y overall survival (OS) was similar among the three groups. In contrast, among patients with KMT2A-MLLT10 (n = 37), those with clonal KRAS-MT (n = 5) had adverse 5-y OS compared with both patients with KRAS-WT (n = 20) (KRAS-WT vs. clonal KRAS-MT: 59.7% vs. 0.0%, P = 0.006) and patients with subclonal KRAS-MT (n = 12) (subclonal KRAS-MT vs. clonal KRAS-MT: 58.3% vs. 0.0%, P = 0.04). According to these results, the effects of the clonality of KRAS-MT on prognosis may depend on which KMT2A fusion is present. DisclosuresNannya: Otsuka Pharmaceutical Co., Ltd.: Consultancy, Speakers Bureau; Astellas: Speakers Bureau. Saito: Toshiba corporation: Research Funding. Ogawa: Kan Research Laboratory, Inc.: Consultancy, Research Funding; Otsuka Pharmaceutical Co., Ltd.: Research Funding; Dainippon-Sumitomo Pharmaceutical, Inc.: Research Funding; Eisai Co., Ltd.: Research Funding; Ashahi Genomics: Current holder of individual stocks in a privately-held company; ChordiaTherapeutics, Inc.: Consultancy, Research Funding.

Persistent Somatic Mutation at Remission Is Associated with Poor Survival and Higher Risk of Relapse in Patients with Acute Myeloid Leukemia

IntroductionMinimal residual disease (MRD) measured by multi-color flow cytometry has been associated with an increased risk of relapse in patients with acute myeloid leukemia (AML). There is also a strong interest in using somatic mutation as an MRD marker and persistence of NPM1 mutation after 2 cycles of induction therapy has been shown to increase the risk of relapse in AML (Ivey et al, NEJM 2016). However, only one third of AML patients carry NPM1 mutation and prognostic impact of persistent mutations in the context of other co-occurring mutations and association with flow cytometry measured MRD is not well understood.MethodsWe studied 95 patients with previously untreated AML who received frontline induction chemotherapy and subsequently achieved complete remission (CR). Paired bone marrow samples obtained at pre-treatment and CR (median day 31) were analyzed by targeted capture exome sequencing of 295 genes (median 309x coverage). Mutect and Pindel was used to call single nucleotide variants (SNVs) and small indels, respectively. We defined two levels of mutation clearance (MC) based on the variant allele frequency (VAF) of the detected mutations at CR: 1) MC2.5, if persistent mutation with VAF <2.5%, and 2) MC1.0 if persistent mutation with VAF <1%. Overall survival (OS), relapse free survival (RFS), event free survival (EFS), and cumulative incidence of relapse (CIR) considering non-relapse death as a competing event were calculated from the date of CR.ResultsOf 95 patients, 49 (52%) were male and 46 (48%) were female. The median age was 53 (range: 17-76). Conventional karyotyping showed one (1%) patient with favorable cytogenetics (CG), 70 (74%) with intermediate CG, and 21 (22%) with adverse CG (all defined by ELN criteria). Eighty nine (94%) patients had de novo AML and 6 (6%) had secondary AML. Thirty-nine (41%) patients underwent allogeneic stem cell transplant in first CR. In the pre-treatment samples of 95 AML patients, we detected 299 high-confidence somatic mutations (192 SNVs and 107 indels) in 60 genes in 85 (89%) patients. The most frequently mutated genes were NPM1 (31%) followed by DNMT3A (29%), FLT3 (18%), and IDH2 (18%). In the matching CR samples, 84 (28%) and 108 (36%) mutations persisted at the level of VAF ≥2.5% and ≥1%, respectively, which corresponded to 41 (48%) and 31 (36%) patients achieving MC2.5 and MC1.0, respectively. Pattern of MC was distinct between mutated genes and affected molecular pathways (Table1). Mutations associated with clonal hematopoiesis of indeterminate potential (CHIP), DNA methylation, and splicing had low rate of MC, whereas mutations in transcription factor genes or receptor tyrosine kinase (RTK) pathway had high rate of MC. With the median follow-up of 41 months (95% CI: 39-47), patients who achieved MC1.0 had significantly better RFS, EFS, OS and lower CIR than those without MC1.0, while there was no significant difference in any of the above outcomes by MC2.5 (Table 2). Flow cytometry-based MRD (flow-MRD) data were available in 92 (97%) patients at CR. Positive flow-MRD was associated with significantly worse RFS, EFS, OS and CIR (Table2). Among 61 patients with negative flow-MRD, 25 (41%) and 32 (52%) had persistent mutations with VAF ≥2.5% and ≥1%, respectively. Multivariable analysis considering mutations that were prognostic in univariable analysis (NPM1, IDH1), age, ELN-defined adverse cytogenetics, flow-MRD and MC revealed that MC1.0 was a significant prognostic factor for RFS (p = 0.03, HR 0.36, 95%CI: 0.14-0.91) and CIR (p = 0.03, HR 0.42, 95%CI: 0.19-0.92). MC1.0 in DNA methylation (2-year EFS: 85% (95% CI: 51-96) in MC1.0 vs. 40% (95% CI: 23-56) in non-MC1.0, p = 0.01; 2-year OS: 85% (95% CI: 51-96) in MC1.0 vs. 58% (95% CI: 39-72) in non-MC1.0, p = 0.03) or in chromatin-cohesin pathways (2-year EFS: 73% (95% CI: 37-90) in MC1.0 vs. 15% (95% CI: 3-39) in non-MC1.0, p < 0.01; 2-year OS: 82% (95% CI: 45-95) in MC1.0 vs. 39% (95% CI: 14-63) in non-MC1.0, p < 0.01) was associated with significantly better EFS and OS.ConclusionIn patients with AML treated with induction chemotherapy, persistent somatic mutation at CR with VAF >1% was associated with increased risk of death and relapse. Assessment of both mutation clearance and flow-based MRD at CR may help further risk stratification of AML. [Display omitted] DisclosuresDaver:Immunogen: Research Funding; Kiromic: Research Funding; Bristol-Myers Squibb Company: Consultancy, Research Funding; Karyopharm: Consultancy, Research Funding; Incyte Corporation: Honoraria, Research Funding; Novartis Pharmaceuticals Corporation: Consultancy; Otsuka America Pharmaceutical, Inc.: Consultancy; Sunesis Pharmaceuticals, Inc.: Consultancy, Research Funding; Daiichi-Sankyo: Research Funding; Jazz: Consultancy; Pfizer Inc.: Consultancy, Research Funding. Cortes:ARIAD: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy, Research Funding; ImmunoGen: Consultancy, Research Funding; Sun Pharma: Research Funding; BMS: Consultancy, Research Funding. Jabbour:Bristol-Myers Squibb: Consultancy. Kantarjian:Bristol-Meyers Squibb: Research Funding; ARIAD: Research Funding; Delta-Fly Pharma: Research Funding; Pfizer: Research Funding; Amgen: Research Funding; Novartis: Research Funding. Takahashi:Symbio Pharmaceuticals: Consultancy.

Outcome of Patients with T-Cell Acute Lymphoblastic Leukemia/Lymphoma with Early T-Cell Precursor-like Immunophenotype with Strong CD5 Expression

Outcome of Patients with T-Cell Acute Lymphoblastic Leukemia/Lymphoma with Early T-Cell Precursor-like Immunophenotype with Strong CD5 Expression

Coexistence and Prognostic Significance of EVI1 Expression and Driver Mutations in KMT2A-Rearranged Acute Myeloid Leukemia

Background: KMT2A(MLL)-rearrangements are among the most frequent chromosomal abnormalities in acute myeloid leukemia (AML) and high EVI1 expression is known as an adverse prognostic factor in this subgroup. A recent study showed driver mutations are associated with poor prognosis in pediatric KMT2A-rearranged AML, however, the relationship between EVI1 expression and driver mutations is unclear. In this study, we examined coexistence and prognostic significance of these genetic abnormalities in pediatric and adult KMT2A-rearranged AML. Patients and Methods: Forty-four pediatric KMT2A-rearranged AML samples collected in the AML-05 study, conducted by the Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG), were analyzed for 338 genes by targeted sequencing. In addition, 85 adult KMT2A-rearranged AML samples in MLL Munich Leukemia Laboratory were analyzed for 16 genes (ASXL1, BRAF, CBL, CEBPA, FLT3-ITD, FLT3-TKD, IDH1, IDH2, KIT, KRAS, NPM1, NRAS, PTPN11, RUNX1, TP53 and WT1) by amplicon deep-sequencing, direct Sanger sequencing, or melting curve analysis. In both studies, EVI1 expression was measured using quantitative RT-PCR. Results and Discussion: In pediatric KMT2A-rearranged AML (n=44), among 28 patients with low EVI1 expression, 13 patients (EVI1-WT, 46.4%) had no driver mutations and 15 patients (EVI1-MT, 53.6%) had one or more driver mutations. Among 16 patients with high EVI1 expression, 2 patients (EVI1+WT, 12.5%) had no driver mutations and 14 patients (EVI1+MT, 87.5%) had one or more driver mutations. Mutations in activated signaling pathway genes (FLT3, NRAS, KRAS, PTPN11, CBL, and BRAF) were detected in most patients with EVI1-MT (12/15, 80.0%) and EVI1+MT (13/14, 93.3%). The frequency of mutations in epigenetic regulator genes (SETD2, ASXL1, ASXL2, BCOR, KDM6A, and CREBBP) was significantly higher in EVI1-MT patients (7/15, 46.7%), compared with EVI1+MT patients (1/14, 7.1%) (P=0.04). By contrast, the frequency of mutations in cohesion complex genes (STAG2 and SMC3) was significantly lower in patients with EVI1-MT patients (0/15, 0.0%), compared with EVI1+MT patients (4/14, 28.6%) (P=0.04). The frequency of mutations in transcription factor genes (WT1, MECOM, SPI1, GATA2, and RUNX1) was also lower in EVI1-MT patients (2/15, 13.3%), compared with EVI1+MT patients (5/14, 35.7%) (P=0.21). In adult patients, the frequency of ASXL1 mutations was higher in EVI1-MT patients (3/18, 18.8%), compared with EVI1+MT patients (1/29, 3.4%) (P=0.15) and those of WT1 and RUNX1 mutations were lower in EVI1-MT patients (0/18, 0.0%), compared with EVI1+MT patients (4/29, 13.8%) (P=0.28), which were compatible with the tendency in pediatric KMT2A-rearranged AML. Next, we examined the prognostic significance of EVI1 expression and driver mutations. Compared with EVI1-WT patients, EVI1-MT patients had lower event-free survival (EFS) (3-years EFS: 84.6% vs. 65.2%, P=0.24). EFS of EVI1+MT patients (3-years EFS: 30.8%) was significantly lower than that of EVI1-WT patients (P=0.001) and EVI1-MT patients (P=0.04). EFS of EVI1+WT patients (3-years EFS: 0.0%) was also lower than that of EVI1-WT patients (P=0.003) and EVI1-MT patients (P=0.09). There was no significant difference in EFS between EVI1+WT and EVI1+MT patients (P=0.88). The results of overall survival (OS) were similar except for EVI1+WT patients (n=2) (3-years OS: EVI1-WT 92.3%, EVI1-MT 70.6%, EVI1+WT 100.0%, EVI1+MT 46.6%). Multivariate analysis including EVI1 expression, driver mutations, age, white blood cell count, and KMT2A-MLLT4 fusion showed EVI1+ is an independent prognostic factor for EFS (hazard ratio (HR): 3.02, 95% confidence interval (CI): 1.08-9.48, P=0.04). There was no prognostic significance in driver mutations (HR:1.24, 95%CI: 0.39-4.74, P=0.73). As a whole, adult patients' survival data were lower, however, the tendency was similar to that of pediatric data (3-years EFS: EVI1-WT 41.7%, EVI1-MT 24.5%, EVI1+WT 9.1%, EVI1+MT 13.5%; 3-years OS: EVI1-WT 55.6%, EVI1-MT 30.7%, EVI1+WT 18.2%, EVI1+MT 36.2%). These data showed that there is an association between EVI1 expression and the pathway of driver mutations, suggesting these abnormalities may have some cooperative mechanisms in leukemogenesis. Compared with driver mutations, high EVI1 expression may have a stronger impact on poor prognosis in KMT2A-rearranged AML, however, the results should be confirmed in the larger cohort. Disclosures Ogawa: RegCell Corporation: Equity Ownership; Kan Research Laboratory, Inc.: Consultancy; ChordiaTherapeutics, Inc.: Consultancy, Equity Ownership; Dainippon-Sumitomo Pharmaceutical, Inc.: Research Funding; Asahi Genomics: Equity Ownership; Qiagen Corporation: Patents &amp; Royalties. Meggendorfer:MLL Munich Leukemia Laboratory: Employment. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Clinical and molecular characterization of patients with acute myeloid leukemia and sole trisomies of chromosomes 4, 8, 11, 13 or 21.

Sole trisomies of chromosomes 4, 8, 11, 13 and 21 account for 89-95% of all sole trisomies in adult AML patients. We analyzed clinical and molecular characteristics of 138 de novo AML patients with sole +4, +8, +11, +13 or +21, and compared them with AML patients with those trisomies occurring in addition to other chromosome abnormalities (non-sole trisomy) and with cytogenetically normal AML (CN-AML) patients. Mutations in methylation-related genes were most commonly observed within each sole trisomy group (+4, 55%; +8, 58%; +11, 71%; +13, 71%; +21, 75% of patients). Patients with sole trisomies, excluding +4, also had frequent mutations in spliceosome genes (+8, 43%; +11, 65%; +13, 65%; +21, 45% of patients). In contrast, +4 patients frequently had mutations in transcription factor genes (44%) and NPM1 (36%). While 48% of patients with sole trisomies harbored mutations in a spliceosome gene, spliceosome mutations were observed in only 24% of non-sole trisomy (n = 131, P < 0.001) and 19% of CN-AML patients (n = 716, P < 0.001). Our data suggest that mutations affecting methylation-related genes are a molecular hallmark of sole trisomies. Mutations in spliceosome genes were also commonly observed in many sole trisomy patients and represent a novel finding in this cytogenetic subgroup.

Hematopoietic transcription factor gene mutations and inherited platelet defects

Hematopoietic transcription factors (TF) are proteins that bind to specific DNA sequences and regulate expression of genes. Hematopoietic TF act in a combinatorial manner to bind sequence-specific DNA within promoter regions to regulate specific gene expression, either as activators or repressors. Hematopoietic TF gene mutations induce rippling downstream effects by simultaneously altering the expression of multiple genes. Mutations involving these hematopoietic TF affect diverse aspects of megakaryocyte biological function, and platelet production and function, culminating in thrombocytopenia and platelet dysfunction. Some hematopoietic TF mutations are associated with predisposition to hematologic malignancies. The molecular and genetic mechanisms in inherited platelet defects (IPD) are unknown. A growing number of evidences suggest that hematopoietic TF gene mutations are important underlying causes for defects in platelet production, morphology and function. The review summarizes the current scientific progress of hematopoietic TF gene mutations in IPD. Key words: Transcription factors; Hematopoietic system; Mutation; Blood platelet disorders; Thrombocytopenia; Inherited platelet defects

A Recurrent Activating Missense Mutation in Waldenström Macroglobulinemia Affects the DNA Binding of the ETS Transcription Factor SPI1 and Enhances Proliferation.

The ETS-domain transcription factors divide into subfamilies based on protein similarities, DNA-binding sequences, and interaction with cofactors. They are regulated by extracellular clues and contribute to cellular processes, including proliferation and transformation. ETS genes are targeted through genomic rearrangements in oncogenesis. The PU.1/SPI1 gene is inactivated by point mutations in human myeloid malignancies. We identified a recurrent somatic mutation (Q226E) in PU.1/SPI1 in Waldenström macroglobulinemia, a B-cell lymphoproliferative disorder. It affects the DNA-binding affinity of the protein and allows the mutant protein to more frequently bind and activate promoter regions with respect to wild-type protein. Mutant SPI1 binding at promoters activates gene sets typically promoted by other ETS factors, resulting in enhanced proliferation and decreased terminal B-cell differentiation in model cell lines and primary samples. In summary, we describe oncogenic subversion of transcription factor function through subtle alteration of DNA binding leading to cellular proliferation and differentiation arrest. SIGNIFICANCE: The demonstration that a somatic point mutation tips the balance of genome-binding pattern provides a mechanistic paradigm for how missense mutations in transcription factor genes may be oncogenic in human tumors.This article is highlighted in the In This Issue feature, p. 681.

Pituitary Transcription Factor Mutations Leading to Hypopituitarism.

Congenital pituitary hormone deficiency is a disabling condition. It is part of a spectrum of disorders including craniofacial midline developmental defects ranging from holoprosencephaly through septo-optic dysplasia to combined and isolated pituitary hormone deficiency. The first genes discovered in the human disease were based on mouse models of dwarfism due to mutations in transcription factor genes. High-throughput DNA sequencing technologies enabled clinicians and researchers to find novel genetic causes of hypopituitarism for the more than three quarters of patients without a known genetic diagnosis to date. Transcription factor (TF) genes are at the forefront of the functional analysis of novel variants of unknown significance due to the relative ease in in vitro testing in a research lab. Genetic testing in hypopituitarism is of high importance to the individual and their family to predict phenotype composition, disease progression and to avoid life-threatening complications such as secondary adrenal insufficiency.This chapter aims to highlight our current understanding about (1) the contribution of TF genes to pituitary development (2) the diversity of inheritance and phenotype features in combined and select isolated pituitary hormone deficiency and (3) provide an initial assessment on how to approach variants of unknown significance in human hypopituitarism. Our better understanding on how transcription factor gene variants lead to hypopituitarism is a meaningful step to plan advanced therapies to specific genetic changes in the future.

Mutaome analysis including 10 common mutated genes in newly diagnosed acute myeloid leukemia patients

Objective To investigate the mutaome profile of the 10 common mutated genes in newly diagnosed acute myeloid leukemia(AML)patients. Methods Gene mutations of ASXL1, CEBPA, DNMT3A, FLT3, IDH1/2, KIT, NPM1, PHF6 and TET2 were analyzed using Sanger sequencing method in bone marrow samples of 129 newly diagnosed AML patients. Results 68.99 %(89/129)patients carried at least one mutation, and 30.23 %(39/129)carried multiple mutations. Mutations of kinase gene FLT3 and KIT were mutually exclusive, and FLT3 mutations occured more often with other mutations, while KIT mutations usually occured alone. Mutations of transcription factor genes CEBPA, NPM1 and PHF6 were accompany with each other. Mutations of epigenetic regulation genes ASXL1, DNMT3A, IDH1/2 and TET2 were mutually exclusive, but were accompany with mutations of kinase or transcription factor genes. Conclusions There are certain rules in the appearance of genetic mutations, and which are related to the function and classification of genes. Key words: Leukemia, myeloid, acute; Newly diagnosed; Gene mutation; Mutaome

Absence of Association Between Length Variation of an Intronic Region in the NFATC1 Gene and Congenital Heart Defects in a Han Chinese Population

Congenital heart defects are complicated birth defects due to the interaction of genetic and environmental factors. Previous research indicated the importance of transcription factors in heart development, which suggested that mutations of transcription factor genes could be genetic determinants of congenital heart defects. Recently, the length variation of an intronic region in the NFATC1 gene was linked to ventricular septal defect (VSD). In this study, we detected the length variation of the region in a Han Chinese population of patients with nonsyndromic VSD, atrial septal defect, patent ductus arteriosus, and control individuals. We found a new allele of the length variation with four repeats of a 44-bp region. At the same time, all the alleles were found in both patient and control groups and there were no significant differences in genotype distribution between the patients and controls. The results suggested no association of the length variation of the intronic region in NFATC1 gene with VSD, atrial septal defect, and patent ductus arteriosus.

Corepressors TLE1 and TLE3 Interact with HESX1 and PROP1

Pituitary hormone deficiency causes short stature in one in 4000 children born and can be caused by mutations in transcription factor genes, including HESX1, PROP1, and POU1F1. HESX1 interacts with a member of the groucho-related gene family, TLE1, through an engrailed homology domain and represses PROP1 activity. Mice with Prop1 deficiency exhibit failed differentiation of the POU1F1 lineage, resulting in lack of TSH, GH, and prolactin. In addition, these mutants exhibit profound pituitary dysmorphology and excess Hesx1 and Tle3 expression. The ability of HESX1 to interact with TLE3 has not been explored previously. We tested the ability of TLE3 to enhance HESX1-mediated repression of PROP1 in cell culture. Both TLE3 and TLE1 repress PROP1 in conjunction with HESX1 with similar efficiencies. TLE1 and TLE3 can each repress PROP1 in the absence of HESX1 via a protein-protein interaction. We tested the functional consequences of ectopic TLE3 and HESX1 expression in transgenic mice by driving constitutive expression in pituitary thyrotrophs and gonadotrophs. Terminal differentiation of these cells was suppressed by HESX1 alone and by TLE3 and HESX1 together but not by TLE3 alone. In summary, we present evidence that HESX1 is a strong repressor that can be augmented by the corepressors TLE1 and TLE3. Our in vitro studies suggest that TLE1 and TLE3 might also play roles independent of HESX1 by interacting with other transcription factors like PROP1.

AML1 mutation and its coexistence with different transcription factor gene families in de novo acute myeloid leukemia: redundancy or synergism

AML1 mutations were identified in 6.3% of AML patients with chromosomal translocations involving CBF, PML-RARalpha, HOX, or ETS transcription factor (TF) gene families. Rare chromosomal abnormalities, t(16;21) and t(7;11), were also found. This study represents the first series to demonstrate the coexistence of known and novel AML1 mutations with different TF gene mutations. Although the occurrence of two TF gene mutations may appear unnecessary, the possible synergistic mechanism between different TF gene families cannot be excluded and needs to be further explored.

Bridging the gap between anatomy and molecular genetics for an improved understanding of congenital heart disease

Birth defects are the leading cause of infant mortality and malformations in congenital heart disease (CHD) are among the most prevalent and fatal of all birth defects. Yet the molecular mechanisms leading to CHD are complex and the causes of the cardiac malformations observed in humans are still unclear. In recent years, the pivotal role of certain transcription factors in heart development has been demonstrated, and gene targeting of cardiac-specific transcription factor genes in animal models has provided valuable insights into heart anomalies. Nonetheless results in these models can be species specific, and in humans, germline mutations in transcription factor genes can only account for some cases of CHD. Furthermore, most patients do not have family history of CHD. There is, therefore, a need for a better understanding of the mechanisms in both normal cardiac development and the formation of malformations. The combining of expertise in cardiac anatomy, pathology, and molecular genetics is essential to adequately comprehend developmental abnormalities associated with CHD. To help elucidate genetic alterations in affected tissues of malformed hearts, we carried out genetic analysis of cardiac-specific transcription factor genes from the Leipzig collection of formalin-fixed malformed hearts. Working with this morphologically well-characterized archival material not only provided valuable genetic information associated with disease, but enabled us to put forward a hypothesis of somatic mutations as a novel molecular cause of CHD. Knowledge of cause and disease mechanism may allow for intervention that could modify the degree of cardiac malformations or development of new approaches for prevention of CHD.

Genetic Analysis of cardiac-specific Transcription Factors Reveals Novel Insights Into Molecular Causes of Congenital Heart Disease

Heart development is complex and requires the sequential and timely interplay of regulatory master proteins, notably several transcription factors. Germline mutations in the human transcription factor genes have been associated with congenital heart disease (CHD), but familial cases studied so far have different mutations. There is no strict genotype-phenotype correlation and mutations in transcription factor genes are rare in unrelated patients. Most cases of CHD come from unaffected family members. The study of archived, but morphologically well-characterized malformed hearts for DNA alterations provides important clues regarding cardiogenic transcription factor genes that would lead to loss-of-function of the protein. Identification of tissue-restricted multiple mutations and multiple haplotypes suggests that somatic mutation and mosaicism are linked to cardiac anomalies. Altogether, somatic mutations and genomic instability in the diseased cardiac tissues of patients with CHD provide a novel mechanism of disease.

Cortical and thalamic axon pathfinding defects in Tbr1, Gbx2, and Pax6 mutant mice: evidence that cortical and thalamic axons interact and guide each other.

During development, cortical areas establish precise reciprocal projections with corresponding thalamic nuclei. Pioneer axons from the cortex and thalamus first meet in the intermediate zone of the subcortical telencephalon (subpallium). Their close interactions in the subpallium suggest that they may use each other for guidance. To test this hypothesis, the development of corticothalamic and thalamocortical connections was studied in mice with mutations of transcription factor genes expressed specifically in the cortex (Tbr1), the dorsal thalamus (Gbx2), or both (Pax6). In Tbr1 mutants, cortical pioneer axons entered the subpallium at the appropriate time, but most stopped growing without entering the diencephalon. Surprisingly, thalamic axons (which do not express Tbr1) deviated into the external capsule and amygdala regions, without entering the cortex. Conversely, in most Gbx2 mutants, thalamic axons were reduced in number and grew no farther than the subpallium. Cortical axons (which do not express Gbx2) grew into the subpallium but did not enter the diencephalon. In one Gbx2- /- case, sparse thalamocortical and corticothalamic projections both developed, but in no case did one projection reach its target and not the other. In Pax6 mutants, neither corticothalamic nor thalamocortical axons reached their targets. These results suggest that thalamocortical and corticothalamic projections may not form independently. After reaching the subpallium, each projection may require a molecularly intact reciprocal projection for further guidance. This type of mechanism ensures that thalamocortical and corticothalamic axons project reciprocally. However, the exact nature of the interaction between cortical and thalamic pioneer axons remains to be elucidated.

Suppression of defective-sporulation phenotypes by mutations in transcription factor genes of Bacillus subtilis

Mutations in the Bacillus subtilis major RNA polymerase sigma factor gene ( rpoD/ crsA47) and a sensory receiver gene ( spoOA/ rvtA11) are potent intergenic suppressors of several stage 0sporulation mutations ( spoOB, OE, OF & OK). We show here that these suppressors also rescue temperature-sensitive sporulation phenotypes (Spo ts) caused by mutations in RNA polymerase, ribosomal protein, and protein synthesis elongation factor EF-G genes. The effects of the crsA and rvtA suppressors on RNA polymerase and ribosomal protein spo ts mutations are similar to those previously described for mutations in another intergenic suppressor gene rev. We have examined the effects of rvtA and crsA mutations on the expression of sporulation-associated membrane proteins, including flagellin and penicillin binding protein 5 ∗ (PBP 5 ∗). Both suppressorsrestored sporulation and synthesis of PBP 5 ∗ in several spoO mutants. However, only rvtA restored flagelli synthesis in spoO suppressed backgrouds. The membrane protein phenotypes resulting from the presence of crsA or rvtA suppressors in spoO strainssuggests that these suppressors function via distinct molecular mechanism. The rvtA and crsA mutations are also able to block the ability of ethanol to induce spoO phenocopies at concentrations of ethanol which prevent sporulation in wild type cells. The effects of ethanol on sporulation-associated membrane protein synthesis in wild type and suppressor containing strains have been examined.