Three Amino Acid Loop Extension Family Research Articles

Comprehensive overview of the role of PBX1 in mammalian kidneys.

Pre-B-cell leukemia homeobox transcription factor 1 (PBX1) is a member of the TALE (three-amino acid loop extension) family and functions as a homeodomain transcription factor (TF). When dimerized with other TALE proteins, it can act as a pioneer factor and provide regulatory sequences via interaction with partners. In vertebrates, PBX1 is expressed during the blastula stage, and its germline variations in humans are interrelated with syndromic anomalies of the kidney, which plays an important role in hematopoiesis and immunity among vertebrates. Herein, we summarize the existing data on PBX1 functions and the impact of PBX1 on renal tumors, PBX1-deficient animal models, and blood vessels in mammalian kidneys. The data indicated that the interaction of PBX1 with different partners such as the HOX genes is responsible for abnormal proliferation and variation of the embryonic mesenchyme, while truncating variants were shown to cause milder phenotypes (mostly cryptorchidism and deafness). Although such interactions have been identified to be the cause of many defects in mammals, some phenotypic variations are yet to be understood. Thus, further research on the TALE family is required.

The TALE never ends: A comprehensive overview of the role of PBX1, a TALE transcription factor, in human developmental defects.

PBX1 is a highly conserved atypical homeodomain transcription factor (TF) belonging to the TALE (three amino acid loop extension) family. Dimerized with other TALE proteins, it can interact with numerous partners and reach dozens of regulating sequences, suggesting its role as a pioneer factor. PBX1 is expressed throughout the embryonic stages (as early as the blastula stage) in vertebrates. In human, PBX1 germline variations are linked to syndromic renal anomalies (CAKUTHED). In this review, we summarized available data on PBX1 functions, PBX1-deficient animal models, and PBX1 germline variations in humans. Two types of genetic alterations were identified in PBX1 gene. PBX1 missense variations generate a severe phenotype including lung hypoplasia, cardiac malformations, and sexual development defects (DSDs). Conversely, truncating variants generate milder phenotypes (mainly cryptorchidism and deafness). We suggest that defects in PBX1 interactions with various partners, including proteins from the HOX (HOXA7, HOXA10, etc.), WNT (WNT9B, WNT3), and Polycomb (BMI1, EED) families are responsible for abnormal proliferation and differentiation of the embryonic mesenchyme. These alterations could explain most of the defects observed in humans. However, some phenotype variability (especially DSDs) remains poorly understood. Further studies are needed to explore the TALE family in greater depth.

Genome-wide Identification, Characterization and Expression profile of TALE gene family in (Juglans regia L.)

The three amino acid loop extension (TALE) are essential transcription factors that regulate the differentiation of meristem in plant growth and development. Several studies on the TALE family in plants, including Arabidopsis, rice, and others, have been carried out. However, little is known about the TALE family in walnut. In this study, the genome-wide identification and expression analysis of the TALE gene family was studied comprehensively in walnut. Based on the specific structure of the TALE domain, a total of thirty walnut TALE genes were identified. According to different domains structure and phylogenetic relationships, the JrTALE gene family could be divided into two subfamilies, namely BELL and KNOX. Conserved protein motifs analysis showed that JrTALE contained similar motif structures in the same phylogenetic branch. The JrTALE gene family members had a weak overall codon bias, preferring to use synonymous codons ending in A/U. Expression pattern analysis of the JrTALE family genes showed that JrTALE17 and JrTALE18 were differentially expressed in female and male flower buds. We confirmed that JrTALE3, JrTALE5, JrTALE7, JrTALE10, JrTALE15 and JrTALE22 were differentially expressed between different stages of flower bud development by real-time quantitative PCR (qRT-PCR). This study provides valuable information about the TALE family in the transitional development of walnut flower buds, laying the foundation for a better understanding of the TALE family in walnut.

Identification and analysis of the TALE transcription factor family in radish

Three-amino acid loop extension (TALE) transcription factors play important roles in plant growth and cell differentiation. There are plenty of studies on TALE transcription factors in several model plants, but not in radish (Raphanus sativas). A genome-wide bioinformatics analysis identified 33 TALE family genes in the Xiang-Ya-Bai (XYB) radish, These genes, are distributed on nine chromosomes and all contain 4-6 exons. The 33 TALE genes in radish showed a co-linearity relationship with the 17 homologous genes in Arabidopsis thaliana. Moreover, a large number of stress response cis-elements were found in the promoter regions of these genes. Expression analysis showed that four genes in the BELL subfamily were highly expressed in roots, and two genes in the KNOX subfamily were highly expressed in shoots of bolting plants and callus. All radish TALE genes contain sequences encoding the conserved HOX domain, except for the gene RSA10037940, which is homologous to Arabidopsis KNATM. The deduced 3D structures of the TALE proteins irrespective of subtypes are highly similar. All the encoded proteins were weakly acidic and hydrophilic. The radish TALE gene family is relatively evolutionarily conserved, which was consistent with results from Arabidopsis, but quite different from that of rice. This study provides important clues for studying the biological functions of TALE transcription factors in radish.

TALE Transcription Factors in Sweet Orange (Citrus sinensis): Genome-Wide Identification, Characterization, and Expression in Response to Biotic and Abiotic Stresses.

Three-amino-acid-loop-extension (TALE) transcription factors comprise one of the largest gene families in plants, in which they contribute to regulation of a wide variety of biological processes, including plant growth and development, as well as governing stress responses. Although sweet orange (Citrus sinensis) is among the most commercially important fruit crops cultivated worldwide, there have been relatively few functional studies on TALE genes in this species. In this study, we investigated 18 CsTALE gene family members with respect to their phylogeny, physicochemical properties, conserved motif/domain sequences, gene structures, chromosomal location, cis-acting regulatory elements, and protein–protein interactions (PPIs). These CsTALE genes were classified into two subfamilies based on sequence homology and phylogenetic analyses, and the classification was equally strongly supported by the highly conserved gene structures and motif/domain compositions. CsTALEs were found to be unevenly distributed on the chromosomes, and duplication analysis revealed that segmental duplication and purifying selection have been major driving force in the evolution of these genes. Expression profile analysis indicated that CsTALE genes exhibit a discernible spatial expression pattern in different tissues and differing expression patterns in response to different biotic/abiotic stresses. Of the 18 CsTALE genes examined, 10 were found to be responsive to high temperature, four to low temperature, eight to salt, and four to wounding. Moreover, the expression of CsTALE3/8/12/16 was induced in response to infection with the fungal pathogen Diaporthe citri and bacterial pathogen Candidatus Liberibacter asiaticus, whereas the expression of CsTALE15/17 was strongly suppressed. The transcriptional activity of CsTALE proteins was also verified in yeast, with yeast two-hybrid assays indicating that CsTALE3/CsTALE8, CsTALE3/CsTALE11, CsTALE10/CsTALE12, CsTALE14/CsTALE8, CsTALE14/CsTALE11 can form respective heterodimers. The findings of this study could lay the foundations for elucidating the biological functions of the TALE family genes in sweet orange and contribute to the breeding of stress-tolerant plants.

Возможная роль гомеодоменсодержащего транскрипционного фактора PREP1 в мезенхимальных клетках сердца

Homeodomain transcription factors play a significant role in mesenchymal stromal cells (MSCs). Previously, the role of Meis1, Pbx1 and Prep1 proteins from the TALE (Three Amino acid Loop Extension) family in adipocytic and osteogenic differentiation of mouse mesenchymal stromal cells was established. In this work, using ChIP-seq and bioinformatic analysis we investigated the binding pattern of PREP1 with the genomic DNA of human heart MSCs, identified nearby genes, and analyzed their ontology. The target genes of the PREP1 factor in cardiac MSCs have been established. Based on the results, the possible involvement of transcription factor PREP1 in the direct reprogramming of fibroblasts into cardiomyocytes is discussed.

Genome-wide Identification and Expression Analysis of TALE Gene Family in Pomegranate (Punica granatum L.)

The three-amino-acid-loop-extension (TALE) gene family is a pivotal transcription factor that regulates the development of flower organs, flower meristem formation, organ morphogenesis and fruit development. A total of 17 genes of pomegranate TALE family were identified and analyzed in pomegranate via bioinformatics methods, which provided a theoretical basis for the functional research and utilization of pomegranate TALE family genes. The results showed that the PgTALE family genes were divided into eight subfamilies (KNOX-Ⅰ, KNOX-Ⅱ, KNOX-Ⅲ, BELL-Ⅰ, BELL-Ⅱ, BELL-Ⅲ, BELL-Ⅳ, and BELL-Ⅴ). All PgTALEs had a KNOX domain or a BELL domain, and their structures were conservative. The 1500 bp promoter sequence had multiple cis-elements in response to hormones (auxin, gibberellin) and abiotic stress, indicating that most of PgTALE were involved in the growth and development of pomegranates and stress. Function prediction and protein-protein network analysis showed that PgTALE may participate in regulating the development of apical meristems, flowers, carpels, and ovules. Analysis of gene expression patterns showed that the pomegranate TALE gene family had a particular tissue expression specificity. In conclusion, the knowledge of the TALE gene gained in pomegranate may be applied to other fruit as well.

Overexpression of Populus transcription factor PtrTALE12 increases axillary shoot development by regulating WUSCHEL expression

The TALE (Three Amino acid Loop Extension) transcription factor family has been shown to control meristem formation and organogenesis in plants. To understand the functional roles of the TALE family in woody perennials, each of the TALE members of Populus trichocarpa was overexpressed in Arabidopsis as a proxy. Among them, the overexpression of PtrTALE12 (i.e., 35S::PtrTALE12) resulted in a dramatic increase of axillary shoot development with early flowering. Interestingly, expression of WUSCHEL (WUS), a central regulator of both apical and axillary meristem formation, was significantly increased in the 35S::PtrTALE12 Arabidopsis plants. Conversely, WUS expression was downregulated in 35S::PtrTALE12-SRDX (short transcriptional repressor domain) plants. Further analysis found that PtrTALE12, expressed preferentially in meristem tissues, directly regulates WUS expression in transient activation assays using Arabidopsis leaf protoplast. Yeast two-hybrid assays showed that PtrTALE12 interacts with SHOOT MERISTEMLESS (STM); however, the interaction does not affect the WUS expression. In addition, expression of both CIRCADIAN CLOCK ASSOCIATED1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY) genes was suppressed accordingly for early flowering 35S::PtrTALE12 Arabidopsis. Indeed, transgenic poplars overexpressing PtrTALE12 as well as Arabidopsis plants overexpressing AtBLH11, a close homolog of PtrTALE12, phenocopied the 35S::PtrTALE12 Arabidopsis (i.e., increased axillary shoot development). Taken together, our results suggest that PtrTALE12 functions as a positive regulator of axillary shoot formation in both Arabidopsis and poplar.

TGIF1 is a negative regulator of MLL-rearranged acute myeloid leukemia.

Members of the TALE (three-amino-acid loop extension) family of atypical homeodomain-containing transcription factors are important downstream effectors of oncogenic fusion proteins involving the mixed lineage leukemia (MLL) gene. A well-characterized member of this protein family is MEIS1, which orchestrates a transcriptional program required for the maintenance of MLL-rearranged acute myeloid leukemia (AML). TGIF1/TGIF2 are relatively uncharacterized TALE transcription factors, which, in contrast to the remaining family, have been shown to act as transcriptional repressors. Given the general importance of this family in malignant hematopoiesis, we therefore tested the potential function of TGIF1 in the maintenance of MLL-rearranged AML. Gene expression analysis of MLL-rearranged patient blasts demonstrated reduced TGIF1 levels, and, in accordance, we find that forced expression of TGIF1 in MLL-AF9-transformed cells promoted differentiation and cell cycle exit in vitro, and delayed leukemic onset in vivo. Mechanistically, we show that TGIF1 interferes with a MEIS1-dependent transcriptional program by associating with MEIS1-bound regions in a competitive manner and that the MEIS1:TGIF1 ratio influence the clinical outcome. Collectively, these findings demonstrate that TALE family members can act both positively and negatively on transcriptional programs responsible for leukemic maintenance and provide novel insights into the regulatory gene expression circuitries in MLL-rearranged AML.

PBX1 is a favorable prognostic biomarker as it modulates 13-cis retinoic acid-mediated differentiation in neuroblastoma.

Neuroblastoma is an embryonic childhood cancer with high mortality. 13-cis retinoic acid (13-cisRA) improves survival for some patients, but many recur, suggesting clinical resistance. The mechanism of resistance and the normal differentiation pathway are poorly understood. Three-amino-acid loop extension (TALE) family genes are master regulators of differentiation. Because retinoids promote differentiation in neuroblastoma, we evaluated TALE family gene expression in neuroblastoma. We evaluated expression of TALE family genes in RA-sensitive and -resistant neuroblastoma cell lines, with and without 13-cisRA treatment, identifying genes whose expression correlates with retinoid sensitivity. We evaluated the roles of one gene, PBX1, in neuroblastoma cell lines, including proliferation and differentiation. We evaluated PBX1 expression in primary human neuroblastoma samples by qRT-PCR, and three independent clinical cohort microarray datasets. We confirmed that induction of PBX1 expression, and no other TALE family genes, was associated with 13-cisRA responsiveness in neuroblastoma cell lines. Exogenous PBX1 expression in neuroblastoma cell lines, mimicking induced PBX1 expression, significantly impaired proliferation and anchorage-independent growth, and promoted RA-dependent and -independent differentiation. Reduced PBX1 protein levels produced an aggressive growth phenotype and RA resistance. PBX1 expression correlated with histologic neuroblastoma subtypes, with highest expression in benign ganglioneuromas and lowest in high-risk neuroblastomas. High PBX1 expression is prognostic of survival, including in multivariate analysis, in the three clinical cohorts. PBX1 is an essential regulator of differentiation in neuroblastoma and potentiates retinoid-induced differentiation. Neuroblastoma cells and tumors with low PBX1 expression have an immature phenotype with poorer prognosis, independent of other risk factors.

Misexpression of Pknox2 in Mouse Limb Bud Mesenchyme Perturbs Zeugopod Development and Deltoid Crest Formation

The TALE (Three Amino acid Loop Extension) family consisting of Meis, Pbx and Pknox proteins is a group of transcriptional co-factors with atypical homeodomains that play pivotal roles in limb development. Compared to the in-depth investigations of Meis and Pbx protein functions, the role of Pknox2 in limb development remains unclear. Here, we showed that Pknox2 was mainly expressed in the zeugopod domain of the murine limb at E10.5 and E11.5. Misexpression of Pknox2 in the limb bud mesenchyme of transgenic mice led to deformities in the zeugopod and forelimb stylopod deltoid crest, but left the autopod and other stylopod skeletons largely intact. These malformations in zeugopod skeletons were recapitulated in mice overexpressing Pknox2 in osteochondroprogenitor cells. Molecular and cellular analyses indicated that the misexpression of Pknox2 in limb bud mesenchyme perturbed the Hox10-11 gene expression profiles, decreased Col2 expression and Bmp/Smad signaling activity in the limb. These results indicated that Pknox2 misexpression affected mesenchymal condensation and early chondrogenic differentiation in the zeugopod skeletons of transgenic embryos, suggesting Pknox2 as a potential regulator of zeugopod and deltoid crest formation.

Dynamic expression of MEIS1 homeoprotein in E14.5 forebrain and differentiated forebrain-derived neural stem cells

Central nervous system development is controlled by highly conserved homeoprotein transcription factors including HOX and TALE (Three Amino acid Loop Extension). TALE proteins are primarily known as HOX-cofactors and play key roles in cell proliferation, differentiation and organogenesis. MEIS1 is a TALE member with established expression in the developing central nervous system. MEIS1 is essential for embryonic development and Meis1 knockout mice dies at embryonic day (E) 14.5. However, Meis1/MEIS1 expression in the devolving forebrain, at this critical time-point has not been studied. Here, for the first time we characterize the region-specific expression of MEIS1 in E14.5 mouse forebrain, filling the gap of MEIS1 expression profile between E12.5 and E16.5. Previously, we reported MEIS1 transcriptional regulatory role in neuronal differentiation and established forebrain-derived neural stem cells (NSC) for gene therapy application of neuronal genes. Here, we show the dynamic expression of Meis1/MEIS1 during the differentiation of forebrain-derived NSC toward a glial lineage. Our results show that Meis1/MEIS1 expression is induced during NSC differentiation and is expressed in both differentiated neurons and astrocytes. Confirming these results, we detected MEIS1 expression in primary cultures of in vivo differentiated cortical neurons and astrocytes. We further demonstrate Meis1/MEIS1 expression relative to other TALE family members in the forebrain-derived NSC in the absence of Hox genes. Our data provide evidence that forebrain-derived NSC can be used as an accessible in vitro model to study the expression and function of TALE proteins, supporting their potential role in modulating NSC self-renewal and differentiation.

PBX1 and MEIS1 up-regulate SOX3 gene expression by direct interaction with a consensus binding site within the basal promoter region

Sox3/SOX3 [SRY (sex determining region Y)-box 3] is considered to be one of the earliest neural markers in vertebrates, playing a role in specifying neuronal fate. We have previously reported characterization of the SOX3 promoter and demonstrated that the general transcription factors NF-Y (nuclear factor-Y), Sp1 (specificity protein 1) and USF (upstream stimulatory factor) are involved in transcriptional regulation of SOX3 promoter activity. In the present study we provide the first evidence that the TALE (three-amino-acid loop extension) transcription factors PBX1 (pre-B-cell leukaemia homeobox 1) and MEIS1 (myeloid ecotropic viral integration site 1 homologue) participate in regulating human SOX3 gene expression in NT2/D1 cells by direct interaction with the consensus PBX/MEIS-binding site, which is conserved in all mammalian orthologue promoters analysed. PBX1 is present in the protein complex formed at this site with nuclear proteins from uninduced cells, whereas both PBX1 and MEIS1 proteins were detected in the complex created with extract from RA (retinoic acid)-induced NT2/D1 cells. By functional analysis we also showed that mutations of the PBX1/MEIS1-binding sites resulted in profound reduction of SOX3 promoter responsiveness to RA. Finally, we demonstrated that overexpressed PBX1 and MEIS1 increased endogenous SOX3 protein expression in both uninduced and RA-induced NT2/D1 cells. With the results of the present study, for the first time, we have established a functional link between the TALE proteins, PBX1 and MEIS1, and expression of the human SOX3 gene. This link is of particular interest since both TALE family members and members of the SOX superfamily are recognized as important developmental regulators.

Cytosolic Sequestration of Prep1 Influences Early Stages of T Cell Development

ObjectivePrep1 and Pbx2 are the main homeodomain DNA-binding proteins of the TALE (three amino acid loop extension) family expressed in the thymus. We previously reported reduced Pbx2 expression and defective thymocyte maturation in Prep1 hypomorphic mice. To further investigate the role of this homeodomain DNA-binding protein in T cell development, we generated transgenic mice expressing the N-terminal fragment of Pbx1 (Pbx1NT) under the control of the Lck proximal promoter.Principal FindingsPbx1NT causes Prep1 cytosolic sequestration, abolishes Prep1-dependent DNA-binding activity and results in reduced Pbx2 expression in developing thymocytes. Transgenic thymi reveal increased numbers of CD4− CD8− CD44− (DN3 and DN4) thymocytes, due to a higher frequency of DN2 and DN4 Pbx1NT thymocytes in the S phase. Transgenic thymocytes however do not accumulate at later stages, as revealed by a normal representation of CD4/CD8 double positive and single positive thymocytes, due to a higher rate of apoptotic cell death of DN4 Pbx1NT thymocytes.ConclusionThe results obtained by genetic (Prep1 hypomorphic) and functional (Pbx1NT transgenic) inactivation of Prep1 support nonredundant roles for this homeodomain protein during different stages of T cell development.

C-Myb Is an Essential Downstream Target for Homeobox Mediated Transformation of Hematopoietic Cells.

Overexpression of Hox genes is frequently associated with leukemogenic transformation of hematopoietic cells. In particular the abdominal-type HoxA genes together with their homeodomain containing dimerization partners of the TALE (three amino acid loop extension) family are well documented oncogenes in various hematologic malignancies. Paradigmatic examples are Hoxa9 and Meis1 that are coactivated by retroviral insertions in the BXH2 mouse leukemia model. In addition Hoxa9 is a fusion partner of NUP98 in acute myeloid leukemias with a t(7;11) and Meis1 as well as Hoxa9 are critical targets of MLL (mixed lineage leukemia) fusion proteins. Despite their important role in leukemogenesis little is known how they exert their transforming function. Here we used a novel system of transient transformation by an inducible oncoprotein to generate populations of primary hematopoietic cells that overexpress Hoxa9 in combination with Meis1. The RNA inventory of these cells was compared to control cells by Affymetrix array analysis. Interestingly the RNA of c-Myb (myeloblastosis leukemia virus oncogene) and of a known c-myb downstream target (Gstm1) was amongst the top ten genes upregulated by Hoxa9/Meis1. c-Myb overexpression was verified by Northern blot and quantitative RT-PCR. c-Myb levels also responded to the introduction of MLL-ENL, an oncoprotein that upregulates Hoxa9 and Meis1. Small interfering RNAs directed against c-Myb suppressed transformation by MLL-ENL but had no effect on transformation of hematopoietic precursors by E2A-HLF an oncogene that does not work by increasing Hoxa9/Meis1 levels. The anti c-Myb siRNA effect was abrogated by coexpression of a c-Myb derivative with a mutated siRNA target site. The coexpression of a dominant negative c-Myb mutant had a similar but weaker effect on MLL-ENL mediated transformation. Ectopic expression of c-Myb in hematopoietic precursor cells induced a mild differentiation block. However, c-Myb alone was not able to give a positive readout in a serial replating assay under conditions where the combined overexpression of Hoxa9 together with Meis1 was clearly transforming. In summary the results suggest that c-Myb is essential but not sufficient for the oncogenic activity of Hoxa9/Meis1.

HOXA10, Pbx2, and Meis1 Protein Expression in the Human Endometrium: Formation of Multimeric Complexes on HOXA10 Target Genes

HOXA10 is a transcription factor necessary for embryonic uterine development and for adult endometrial receptivity. The three-amino acid loop extension family of cofactors, including Pbx and Meis, provide HOX target gene specificity in development and myeloid differentiation. Here we demonstrate the expression of Pbx and Meis family cofactors in the human endometrium and their interaction with HOXA10. Using immunohistochemical analysis, we found that Pbx2 and Meis1, but not Pbx1, Pbx3, or Meis2, were expressed in human endometrium. HOXA10, Pbx2, and Meis1 were expressed in the stroma throughout the menstrual cycle. The glandular expression of HOXA10 and Meis1 was menstrual cycle stage specific, whereas glandular Pbx2 expression did not vary. Pbx2, but not Meis1, was expressed in Ishikawa cells. EMSA demonstrated HOXA10-Pbx2 binding as a heterodimer to an enhancer of the EMX2 gene, a known target of HOXA10 regulation. Ablation of the Pbx binding site, but not ablation of the HOXA10 binding site in EMX2, resulted in loss of dimer binding. Based on the observed expression and binding patterns of Pbx2, Meis1, and HOXA10, it is likely that heterodimeric and trimeric complexes involving these proteins determine HOXA10 target gene specificity. Enhanced target gene specificity imparted by multimer binding is likely necessary for HOXA10-mediated endometrial receptivity.