Loop Helix Transcription Factor Research Articles

Mechanical stress mediated by both endosperm softening and embryo growth underlies endosperm elimination in Arabidopsis seeds.

Seed development in angiosperms demands the tightly coordinated development of three genetically distinct structures. The embryo is surrounded by the endosperm, which is in turn enclosed within the maternally derived seed coat. In Arabidopsis, final seed size is determined by early expansion of the coenocytic endosperm, which then cellularises and subsequently undergoes developmental programmed cell death, breaking down as the embryo grows. Endosperm breakdown requires the endosperm-specific basic helix-loop-helix transcription factor ZHOUPI. However, to date, the mechanism underlying the Arabidopsis endosperm breakdown process has not been elucidated. Here, we provide evidence that ZHOUPI does not induce the developmental programmed cell death of the endosperm directly. Instead ZHOUPI indirectly triggers cell death by regulating the expression of cell wall-modifying enzymes, thus altering the physical properties of the endosperm to condition a mechanical environment permitting the compression of the cellularised endosperm by the developing embryo.

Interacting mechanism of ID3 HLH domain towards E2A/E12 transcription factor – An Insight through molecular dynamics and docking approach

Inhibitor of DNA binding protein 3 (ID3) has long been characterized as an oncogene that implicates its functional role through its Helix–Loop–Helix (HLH) domain upon protein–protein interaction. An insight into the dimerization brought by this domain helps in identifying the key residues that favor the mechanism behind it. Molecular dynamics (MD) simulations were performed for the HLH proteins ID3 and Transcription factor E2-alpha (E2A/E12) and their ensemble complex (ID3-E2A/E12) to gather information about the HLH domain region and its role in the interaction process. Further evaluation of the results by Principal Component Analysis (PCA) and Free Energy Landscape (FEL) helped in revealing residues of E2A/E12: Lys570, Ala595, Val598, and Ile599 and ID3: Glu53, Gln63, and Gln66 buried in their HLH motifs imparting key roles in dimerization process. Furthermore the T-pad analysis results helped in identifying the key fluctuations and conformational transitions using the intrinsic properties of the residues present in the domain region of the proteins thus specifying their crucial role towards molecular recognition. The study provides an insight into the interacting mechanism of the ID3-E2A/E12 complex and maps the structural transitions arising in the essential conformational space indicating the key structural changes within the helical regions of the motif. It thereby describes how the internal dynamics of the proteins might regulate their intrinsic structural features and its subsequent functionality.

ZmZHOUPI, an endosperm‐specific basic helix–loop–helix transcription factor involved in maize seed development

In angiosperm seeds the embryo is embedded within the endosperm, which is in turn enveloped by the seed coat, making inter-compartmental communication essential for coordinated seed growth. In this context the basic helix-loop-helix domain transcription factor AtZHOUPI (AtZOU) fulfils a key role in both the lysis of the transient endosperm and in embryo cuticle formation in Arabidopsis thaliana. In maize (Zea mays), a cereal with a persistent endosperm, a single gene, ZmZOU, falls into the same phylogenetic clade as AtZOU. Its expression is limited to the endosperm where it peaks during the filling stage. In ZmZOU-RNA interference knock-down lines embryo size is slightly reduced and the embryonic suspensor and the adjacent embryo surrounding region show retarded breakdown. Ectopic expression of ZmZOU reduces stomatal number, possibly due to inappropriate protein interactions. ZmZOU forms functional heterodimers with AtICE/AtSCREAM and the closely related maize proteins ZmICEb and ZmICEc, but its interaction is more efficient with the ZmICEa protein, which shows sequence divergence and only has close homologues in other monocotyledonous species. Consistent with the observation that these complexes can trans-activate target gene promoters from Arabidopsis, ZmZOU partially complements the Atzou-4 mutant. However, structural, trans-activation and gene expression data support the hypothesis that ZmZOU and ZmICEa may have coevolved to form a functional complex unique to monocot seeds. This divergence may explain the reduced functionality of ZmZOU in Arabidopsis, and reflect functional specificities which are unique to the monocotyledon lineage.

Anthocyanin profile characterization and quantitative trait locus mapping in zicaitai (Brassica rapa L. ssp. chinensis var. purpurea)

Anthocyanins have several biological functions in plants and are beneficial to human health. To elucidate the metabolic profile of anthocyanins and determine the genetic basis controlling anthocyanin accumulation in zicaitai (Brassica rapa L. ssp. chinensis var. purpurea), we conducted anthocyanin profile characterization and quantitative trait locus (QTL) analysis. Seventeen anthocyanin compounds were identified as cyanidin glycosides in zicaitai. A genetic linkage map based on 200 F2 lines was constructed using 161 insertion/deletion markers. Total anthocyanin content (TAC) was determined by pH differential spectrophotometry for the F2 lines. Using the map and phenotypic data, a major QTL which explained 56.7 % of phenotypic variation was identified for TAC on chromosome A09. Two genes, BrEGL3.1 and BrEGL3.2, as syntenic orthologs of AtEGL3 encoding basic helix–loop–helix transcription factors in this QTL region, are candidate genes for a key role in the control of anthocyanin accumulation in zicaitai.

Differential levels of Neurod establish zebrafish endocrine pancreas cell fates

During development a network of transcription factors functions to differentiate foregut cells into pancreatic endocrine cells. Differentiation of appropriate numbers of each hormone-expressing endocrine cell type is essential for the normal development of the pancreas and ultimately for effective maintenance of blood glucose levels. A fuller understanding of the details of endocrine cell differentiation may contribute to development of cell replacement therapies to treat diabetes. In this study, by using morpholino and gRNA/Cas9 mediated knockdown we establish that differential levels of the basic-helix loop helix (bHLH) transcription factor Neurod are required for the differentiation of distinct endocrine cell types in developing zebrafish. While Neurod plays a role in the differentiation of all endocrine cells, we find that differentiation of glucagon-expressing alpha cells is disrupted by a minor reduction in Neurod levels, whereas differentiation of insulin-expressing beta cells is less sensitive to Neurod depletion. The endocrine cells that arise during embryonic stages to produce the primary islet, and those that arise subsequently during larval stages from the intra-pancreatic duct (IPD) to ultimately contribute to the secondary islets, show similar dependence on differential Neurod levels. Intriguingly, Neurod-deficiency triggers premature formation of endocrine precursors from the IPD during early larval stages. However, the Neurod-deficient endocrine precursors fail to differentiate appropriately, and the larvae are unable to maintain normal glucose levels. In summary, differential levels of Neurod are required to generate endocrine pancreas subtypes from precursors during both embryonic and larval stages, and Neurod function is in turn critical to endocrine function.

Expression of the Olig gene family in the developing mouse inner ear

Transcription factors are believed to play key roles in determining cell fate in inner ear development. Olig genes, which are basic helix–loop–helix transcription factors, have been reported to play important roles in the development of the central nervous system. However, members of this family have not previously been implicated in inner ear development, despite the similarity between otocyst and neural tube development. Olig1 begins to be expressed at the ventral domain of the otocyst at embryonic day (E) 9.5, and Olig1 expression in the epithelium of the developing inner ear persists to E15.5. Olig2 expression is localized to the cochleovestibular ganglia from E12.5 through E14.5. Olig3 has a diffuse expression pattern in the developing inner ear from E12.5 through the postnatal stage. Furthermore, at early stages of inner ear development, the Olig1 expression domain overlaps a region that is positive for Sox2 and Jagged1. This observation indicates that Olig1 may play an important role in the specification of the prosensory domain in the developing inner ear. As Olig genes are expressed in the mouse developing inner ear in a temporospatially distinct fashion, they may play substantial roles in the regulation of mammalian inner ear development.

Requirement of the transcription factor USF1 in bovine oocyte and early embryonic development.

Upstream stimulating factor 1 (USF1) is a basic helix-loop-helix transcription factor that specifically binds to E-box DNA motifs, known cis-elements of key oocyte expressed genes essential for oocyte and early embryonic development. However, the functional and regulatory role of USF1 in bovine oocyte and embryo development is not understood. In this study, we demonstrated that USF1 mRNA is maternal in origin and expressed in a stage specific manner during the course of oocyte maturation and preimplantation embryonic development. Immunocytochemical analysis showed detectable USF1 protein during oocyte maturation and early embryonic development with increased abundance at 8-16-cell stage of embryo development, suggesting a potential role in embryonic genome activation. Knockdown of USF1 in germinal vesicle stage oocytes did not affect meiotic maturation or cumulus expansion, but caused significant changes in mRNA abundance for genes associated with oocyte developmental competence. Furthermore, siRNA-mediated depletion of USF1 in presumptive zygote stage embryos demonstrated that USF1 is required for early embryonic development to the blastocyst stage. A similar (USF2) yet unique (TWIST2) expression pattern during oocyte and early embryonic development for related E-box binding transcription factors known to cooperatively bind USF1 implies a potential link to USF1 action. This study demonstrates that USF1 is a maternally derived transcription factor required for bovine early embryonic development, which also functions in regulation of JY1, GDF9, and FST genes associated with oocyte competence.

The circadian clock machinery controls adiponectin expression

Adiponectin, an adipokine involved in glucose and lipid metabolism, exhibits a circadian manner of expression. Adiponectin expression is mediated by the helix–loop–helix transcription factor sterol regulatory element binding protein (SREBP)-1c. In this study, we tested whether the circadian clock helix–loop–helix transcription factors CLOCK and BMAL1 regulate adiponectin expression. We found that adiponectin expression is regulated by the clock through the circadian expression of its transcription factor peroxisome proliferator-activated receptor γ (PPARγ) and its co-activator PPARγ co-activator 1α (PGC1α) in mouse white adipose tissue and differentiated adipocytes. In addition, reconstitution of the core clock mechanism and siRNA experiments in cell culture suggest that the clock directly activates the adiponectin promoter and mediates its expression. In summary, adiponectin expression is regulated by the circadian clock and through the circadian expression of its transcription factor PPARγ and its co-activator PGC1α.

Shh regulates chick Ebf1 gene expression in somite development

The chick early B-cell factor 1 (cEbf1) is a member of EBF family of helix loop helix transcription factors. Recently, we have proved that cEbf1 expression in feather is regulated by Shh. It is therefore possible that the somitic expression of cEbf1 is controlled by Shh signals from the notochord. To assess this hypothesis, the expression profile of cEbf1 was first detailed in somites of chick embryos (from HH8 to HH28). cEbf1 expression was mainly localised in the medial sclerotome and later around the vertebral cartilage anlagen of body and pedicles. Tissue manipulations (notochord ablation) and Shh gain and loss of function experiments were then performed to analyse whether the notochord and/or Shh regulate cEbf1 expression. Results from these experiments confirmed our hypothesis that the medial somitic expression of cEbf1 is regulated by Shh from the notochord. In conclusion, cEbf1 gene is considered as a medial sclerotome marker, downstream to and regulated by the notochord derived Shh, which may be functionally involved in somitogenesis.

Abstract 5458: Methylene blue selectively inhibits B cell development and E2A activity

Abstract Introduction: E2A, a basic helix loop helix transcription factor, is essential for B lymphocyte proliferation, survival and differentiation. Mutations in E2A are associated with B cell leukemia and lymphomas. Our preliminary data suggests that E2A activity is altered by HSP70 and since HSP70 ATPase activity is inhibited by methylene blue (MB), we hypothesized that MB will alter E2A activity and thus alter B cell development. Methods: We studied the effects of MB on normal and leukemic B cells compared to its effect on myeloid cells. The cell lines used were REH, NALM-6 and BAF3-p185 cells compared to myeloid cell lines, CCRF and CMK for proliferation assay. To study MB effect on normal B and myeloid progenitors methylcellulose colony formation assays with murine bone marrow cells (strain 129S1/SVImJ, 8-12 weeks old mice). We also treated mice with MB in their drinking water (1000 μM) up to 16 weeks to study the in-vivo effect on B cells as compared to granulocyte, macrophages, T and NK cells in the peripheral blood by flow analysis. To understand the mechanisms by which MB alters B cell proliferation, we examined the effects of MB on cell cycle progression, apoptosis and E2A activity in B cells. For cell progression NALM-6 cells were treated from 0, 6, 12 and 24 hrs and assayed using BRDU staining by flow analysis. Apoptosis was measured using Annexin V staining by flow analysis in NALM-6 cells up to 48hrs after treatment with MB. RNA isolated from NALM-6 cells treated up to 24hrs was used Prime View gene array analysis. E2A activity was measured using an E2A-specific luciferase reporter assay in 293T cells. Results: MB decreased the proliferation of precursor B cell lines, REH, NALM-6 and BAF3-p185 cells compared to myeloid cell lines, CCRF and CMK. Similar effects of MB were also seen with normal B and myeloid progenitors as assayed by methylcellulose colony formation assays with murine bone marrow cells. Similarly there is a significant decrease in peripheral B cells as compared to granulocytes, macrophages, T and NK cells when the mice were treated with 1000 μM of methylene blue in their drinking water as compared to mice that remained untreated. MB caused apoptosis and decreased the S phase fraction of treated NALM-6 cells. Genes belonging to positive regulation of apoptosis, JNK and p53 pathways were up regulated at both 12 and 24 hrs after treatment with MB. We also found that MB inhibited E2A activity using an E2A-specific luciferase reporter assay. Conclusion: Our findings suggest that MB is a specific inhibitor of B cell proliferation, survival and development. This may be mediated through apoptosis and by inhibition of E2A activity by methylene blue. Citation Format: Eroica Soans, John Kim Choi. Methylene blue selectively inhibits B cell development and E2A activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5458. doi:10.1158/1538-7445.AM2014-5458

Tomato Male sterile 1035 is essential for pollen development and meiosis in anthers.

Male fertility in flowering plants depends on proper cellular differentiation in anthers. Meiosis and tapetum development are particularly important processes in pollen production. In this study, we showed that the tomato male sterile (ms10(35)) mutant of cultivated tomato (Solanum lycopersicum) exhibited dysfunctional meiosis and an abnormal tapetum during anther development, resulting in no pollen production. We demonstrated that Ms10(35) encodes a basic helix-loop-helix transcription factor that is specifically expressed in meiocyte and tapetal tissue from pre-meiotic to tetrad stages. Transgenic expression of the Ms10(35) gene from its native promoter complemented the male sterility of the ms10(35) mutant. In addition, RNA-sequencing-based transcriptome analysis revealed that Ms10(35) regulates 246 genes involved in anther development processes such as meiosis, tapetum development, cell-wall degradation, pollen wall formation, transport, and lipid metabolism. Our results indicate that Ms10(35) plays key roles in regulating both meiosis and programmed cell death of the tapetum during microsporogenesis.

Pdx1 and USF transcription factors co-ordinately regulate Alx3 gene expression in pancreatic β-cells.

Alterations in transcription factors expressed in insulin-producing islet β-cells generate pancreatic dysfunction leading to diabetes. The homeodomain transcription factor Alx3 (aristaless-like homeobox 3) expressed in pancreatic islets participates in the regulated expression of several islet genes, and its deficiency in mice leads to islet cell apoptosis and glucose intolerance. In the present study, we investigated the mechanisms that regulate expression of Alx3in pancreatic islets at the transcriptional level. We found that the Alx3 promoter contains at least eight putative regulatory elements with an E-box consensus sequence, three of which were determined to be functional and required for Alx3 promoter activity by mutational analysis in transfected MIN6 β-cells. We determined that these E-box elements are recognized by the basic helix-loop-helix transcription factors USF1 (upstream stimulatory factor 1) and USF2. We also identified a highly conserved A-box in the Alx3 promoter that is recognized by the islet-specific transcription factor Pdx1 (pancreatic and duodenal homeobox 1). Pdx1-mediated transactivation of the Alx3 promoter requires the integrity of the three functional E-boxes and the co-operation with USF transcription factors bound to them. The results from the present study indicate that Pdx1 contributes to the transcriptional transactivation of Alx3in pancreatic β-cells by acting in co-ordination with USF1 and USF2.

Basic helix-loop-helix transcription factor BcbHLHpol functions as a positive regulator of pollen development in non-heading Chinese cabbage.

Cytoplasmic male sterility (CMS) is a common trait in higher plants, and several transcription factors regulate pollen development. Previously, we obtained a basic helix-loop-helix transcription factor, BcbHLHpol, via suppression subtractive hybridization in non-heading Chinese cabbage. However, the regulatory function of BcbHLHpol during anther and pollen development remains unclear. In this study, BcbHLHpol was cloned, and its tissue-specific expression profile was analyzed. The results of real-time polymerase chain reaction showed that BcbHLHpol was highly expressed in maintainer buds and that the transcripts of BcbHLHpol significantly decreased in the buds of pol CMS. A virus-induced gene silencing vector that targets BcbHLHpol was constructed and transformed into Brassica campestris plants to further explore the function of BcbHLHpol. Male sterility and short stature were observed in BcbHLHpol-silenced plants. The degradation of tapetal cells was inhibited in BcbHLHpol-silenced plants, and nutrients were insufficiently supplied to the microspore. These phenomena resulted in pollen abortion. This result indicates that BcbHLHpol functions as a positive regulator in pollen development. Yeast two-hybrid and bimolecular fluorescence complementation assays revealed that BcbHLHpol interacted with BcSKP1 in the nucleus. This finding suggests that BcbHLHpol and BcSKP1 are positive coordinating regulators of pollen development. Quantitative real-time PCR indicated that BcbHLHpol and BcSKP1 can be induced at low temperatures. Thus, we propose that BcbHLHpol is necessary for meiosis. This study provides insights into the regulatory functions of the BcbHLHpol network during anther development.

Differential responsiveness of distinct retinal domains to Atoh7

During vertebrate eye development retinal progenitor cells (RPCs) differentiate into all neural cell types of the retina. Retinal ganglion cells (RGCs) represent the first cell type to be generated. For their development, Atoh7, a basic Helix Loop Helix (bHLH) transcription factor is crucial. Atoh7 loss of function results in a massive reduction or even a total loss of RGCs. However, inconsistent results have been obtained in atoh7 gain of function experiments with respect to ganglion cell genesis, implying that the effect of Atoh7 is likely to be dependent on the competence state of the RPC.In this study we addressed the differential susceptibilities of early RPCs to Atoh7 in vivo, using medaka. Unexpectedly, we observed a largely normal development of the dorsal retina, although atoh7 was precociously expressed. However, the development of the retina close to the optic nerve head (part of the ventral retina) was disturbed severely. Photoreceptors were largely absent and the Müller glia cell number was reduced significantly. The majority of cells in this domain were ganglion cells and the abnormal development of this area affected the closure of the optic fissure resulting in coloboma.

Nicotine-mediated invasion and migration of non-small cell lung carcinoma cells by modulating STMN3 and GSPT1 genes in an ID1-dependent manner.

BackgroundInhibitor of DNA binding/Differentiation 1 (ID1) is a helix loop helix transcription factor that lacks the basic DNA binding domain. Over-expression of ID1 has been correlated with a variety of human cancers; our earlier studies had shown that reported ID1 is induced by nicotine or EGF stimulation of non-small cell lung cancer (NSCLC) cells and its down regulation abrogates cell proliferation, invasion and migration. Here we made attempts to identify downstream targets of ID1 that mediate these effects.MethodsA microarray analysis was done on two different NSCLC cell lines (A549 and H1650) that were transfected with a siRNA to ID1 or a control, non-targeting siRNA. Cells were stimulated with nicotine and genes that were differentially expressed upon nicotine stimulation and ID1 depletion were analyzed to identify potential downstream targets of ID1. The prospective role of the identified genes was validated by RT-PCR. Additional functional assays were conducted to assess the role of these genes in nicotine induced proliferation, invasion and migration. Experiments were also conducted to elucidate the role of ID1, which does not bind to DNA directly, affects the expression of these genes at transcriptional level.ResultsA microarray analysis showed multiple genes are affected by the depletion of ID1; we focused on two of them: Stathmin-like3 (STMN3), a microtubule destabilizing protein, and GSPT1, a protein involved in translation termination; these proteins were induced by both nicotine and EGF in an ID1 dependent fashion. Overexpression of ID1 in two different cell lines induced STMN3 and GSPT1 at the transcriptional level, while depletion of ID1 reduced their expression. STMN3 and GSPT1 were found to facilitate the proliferation, invasion and migration of NSCLC cells in response to nAChR activation. Attempts made to assess how ID1, which is a transcriptional repressor, induces these genes showed that ID1 down regulates the expression of two transcriptional co-repressors, NRSF and ZBP89, involved in the repression of these genes.ConclusionsCollectively, our data suggests that nicotine and EGF induce genes such as STMN3 and GSPT1 to promote the proliferation, invasion and migration of NSCLC, thus enhancing their tumorigenic properties. These studies thus reveal a central role for ID1 and its downstream targets in facilitating lung cancer progression.

Generation of induced neurons by direct reprogramming in the mammalian cochlea

Primary auditory neurons (ANs) in the mammalian cochlea play a critical role in hearing as they transmit auditory information in the form of electrical signals from mechanosensory cochlear hair cells in the inner ear to the brainstem. Their progressive degeneration is associated with disease conditions, excessive noise exposure and aging. Replacement of ANs, which lack the ability to regenerate spontaneously, would have a significant impact on research and advancement in cochlear implants in addition to the amelioration of hearing impairment. The aim of this study was to induce a neuronal phenotype in endogenous non-neural cells in the cochlea, which is the essential organ of hearing. Overexpression of a neurogenic basic helix–loop–helix transcription factor, Ascl1, in the cochlear non-sensory epithelial cells induced neurons at high efficiency at embryonic, postnatal and juvenile stages. Moreover, induced neurons showed typical properties of neuron morphology, gene expression and electrophysiology. Our data indicate that Ascl1 alone or Ascl1 and NeuroD1 is sufficient to reprogram cochlear non-sensory epithelial cells into functional neurons. Generation of neurons from non-neural cells in the cochlea is an important step for the regeneration of ANs in the mature mammalian cochlea.

Genetic analysis reveals a novel locus in Vicia faba decoupling pigmentation in the flower from that in the extra-floral nectaries

The genetics of the stipule spot pigmentation (SSP) in faba bean (Vicia faba L.) was studied using four inbred lines, of which Disco/2 was zero-tannin (zt2) with colourless stipule spots, ILB938/2 was normal-tannin (ZT2) with colourless stipule spots, and both Aurora/2 and Melodie/2 were ZT2 with coloured stipule spots. Crosses Melodie/2 × ILB 938/2, Melodie/2 × Disco/2, ILB 938/2 × Aurora/2 and ILB 938/2 × Disco/2 (A, B, C and D, respectively) were prepared, along with reciprocals and backcrosses, and advanced through single-seed descent. All F1 hybrid plants had pigmented stipule spots, and in the F2 generation, the segregation ratio fit 3 coloured:1 colourless in crosses A, B and C and 9:7 in cross D. In the F3 generation, the ratio fit 5:3 in crosses A and C and 25:39 in cross D, and in the F4 generation, 9:7 in cross A. SSP was linked to the zero-tannin characteristics (white flower) only in cross B. The results show that coloured stipule spot is dominant to colourless and that colouration is determined by two unlinked complementary recessive genes. We propose the symbols ssp2 for the gene associated with zt2 in Disco/2 and ssp1 for the gene not associated with tannin content in ILB938/2. The novel ssp1 locus was mapped at F5 in cross ‘A’ using Medicago truncatula-derived single-nucleotide polymorphism and was on chromosome 1 of faba bean, in a well-conserved region of M. truncatula chromosome 5 containing some candidate Myb and basic helix–loop–helix transcription factor genes.

MiR396a-Mediated basic helix-loop-helix transcription factor bHLH74 repression acts as a regulator for root growth in Arabidopsis seedlings.

miR396 targets seven GROWTH-REGULATING FACTOR (GRF) genes and the BASIC HELIX-LOOP-HELIX (bHLH) TRANSCRIPTION FACTOR 74 gene (bHLH74) in Arabidopsis. Previous research revealed that the miR396 target module regulates cell proliferation and plays a critical role in leaf development. However, no additional biological functions of miR396 have been investigated in detail. In this study, T-DNA insertion mutants and transgenic plants with altered levels of miR396 or its target genes were used to characterize the regulatory role of miR396 in root development. We found that AtMIR396a was the predominant source for miR396 accumulation in the roots of seedlings, and that the mir396a-1 mutant had longer roots than wild-type seedlings. Overexpression of AtMIR396a decreased the transcript levels of target genes such as GRF genes and bHLH74, and resulted in a shorter root phenotype. Furthermore, the bhlh74-1 mutant had shorter roots, whereas overexpression of an miR396-resistant form of bHLH74 (mbHLH74) had an enhanced root growth phenotype. Moreover, MIR396a regulated root growth by affecting the elongation zone. Taken together, these data indicate that miR396a-mediated bHLH74 repression helps regulate root growth in Arabidopsis seedlings.

Early B-cell factor 1 is an essential transcription factor for postnatal glomerular maturation

The coordination of multiple cytokines and transcription factors with their downstream signaling pathways have been shown to be integral to nephron maturation. Here we present a completely novel role for the helix-loop-helix transcription factor Early B cell Factor 1 (Ebf1), originally identified for B cell maturation, for the proper maturation of glomerular cells from mesenchymal progenitors. The expression of Ebf1 was both spatially and temporally regulated within the developing cortex and glomeruli. Using Ebf1-null mice we then identified biochemical, metabolic, and histological abnormalities in renal development that arose in the absence of this transcription factor. In the Ebf1 knockout mice the developed kidneys show thinned cortices and reduced glomerular maturation. The glomeruli showed abnormal vascularization and severely effaced podocytes. The mice exhibited early albuminuria and elevated blood urea nitrogen levels. Moreover, the GFR was reduced over 66 percent and the expression of podocyte-derived VEGF-A was decreased compared to wild type control mice. Thus, Ebf1 has a significant and novel role in glomerular development, podocyte maturation, and the maintenance of kidney integrity and function.

The bHLH Factors Extramacrochaetae and Daughterless Control Cell Cycle in Drosophila Imaginal Discs through the Transcriptional Regulation of the cdc25 Phosphatase string

One of the major issues in developmental biology is about having a better understanding of the mechanisms that regulate organ growth. Identifying these mechanisms is essential to understand the development processes that occur both in physiological and pathological conditions, such as cancer. The E protein family of basic helix-loop helix (bHLH) transcription factors, and their inhibitors the Id proteins, regulate cell proliferation in metazoans. This notion is further supported because the activity of these factors is frequently deregulated in cancerous cells. The E protein orthologue Daughterless (Da) and the Id orthologue Extramacrochaetae (Emc) are the only members of these classes of bHLH proteins in Drosophila. Although these factors are involved in controlling proliferation, the mechanism underlying this regulatory activity is poorly understood. Through a genetic analysis, we show that during the development of epithelial cells in the imaginal discs, the G2/M transition, and hence cell proliferation, is controlled by Emc via Da. In eukaryotic cells, the main activator of this transition is the Cdc25 phosphatase, string. Our genetic analyses reveal that the ectopic expression of string in cells with reduced levels of Emc or high levels of Da is sufficient to rescue the proliferative defects seen in these mutant cells. Moreover, we present evidence demonstrating a role of Da as a transcriptional repressor of string. Taken together, these findings define a mechanism through which Emc controls cell proliferation by regulating the activity of Da, which transcriptionally represses string.