scholarly journals Basic Helix-Loop-Helix Transcription Factor Heterocomplex of Yas1p and Yas2p Regulates Cytochrome P450 Expression in Response to Alkanes in the Yeast Yarrowia lipolytica

2007 ◽  
Vol 6 (4) ◽  
pp. 734-743 ◽  
Author(s):  
Setsu Endoh-Yamagami ◽  
Kiyoshi Hirakawa ◽  
Daisuke Morioka ◽  
Ryouichi Fukuda ◽  
Akinori Ohta
Keyword(s):  
Cytochrome P450 ◽  
Transcription Factors ◽  
Yarrowia Lipolytica ◽  
Bhlh Protein ◽  
Genome Database ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Bhlh Transcription Factors

ABSTRACT The expression of the ALK1 gene, which encodes cytochrome P450, catalyzing the first step of alkane oxidation in the alkane-assimilating yeast Yarrowia lipolytica, is highly regulated and can be induced by alkanes. Previously, we identified a cis-acting element (alkane-responsive element 1 [ARE1]) in the ALK1 promoter. We showed that a basic helix-loop-helix (bHLH) protein, Yas1p, binds to ARE1 in vivo and mediates alkane-dependent transcription induction. Yas1p, however, does not bind to ARE1 by itself in vitro, suggesting that Yas1p requires another bHLH protein partner for its DNA binding, as many bHLH transcription factors function by forming heterodimers. To identify such a binding partner of Yas1p, here we screened open reading frames encoding proteins with the bHLH motif from the Y. lipolytica genome database and identified the YAS2 gene. The deletion of the YAS2 gene abolished the alkane-responsive induction of ALK1 transcription and the growth of the yeast on alkanes. We revealed that Yas2p has transactivation activity. Furthermore, Yas1p and Yas2p formed a protein complex that was required for the binding of these proteins to ARE1. These findings allow us to postulate a model in which bHLH transcription factors Yas1p and Yas2p form a heterocomplex and mediate the transcription induction in response to alkanes.

scholarly journals Calcium Regulation of Myogenesis by Differential Calmodulin Inhibition of Basic Helix-Loop-Helix Transcription Factors

2008 ◽  
Vol 19 (6) ◽  
pp. 2509-2519 ◽  
Author(s):  
Jannek Hauser ◽  
Juha Saarikettu ◽  
Thomas Grundström
Keyword(s):  
Transcription Factors ◽  
Target Genes ◽  
Myogenic Differentiation ◽  
Bhlh Protein ◽  
Channel Blockers ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Protein ◽  
Bhlh Transcription Factors

The members of the MyoD family of basic helix-loop-helix (bHLH) transcription factors are critical regulators of skeletal muscle differentiation that function as heterodimers with ubiquitously expressed E-protein bHLH transcription factors. These heterodimers must compete successfully with homodimers of E12 and other E-proteins to enable myogenesis. Here, we show that E12 mutants resistant to Ca2+-loaded calmodulin (CaM) inhibit MyoD-initiated myogenic conversion of transfected fibroblasts. Ca2+ channel blockers reduce, and Ca2+ stimulation increases, transcription by coexpressed MyoD and wild-type E12 but not CaM-resistant mutant E12. Furthermore, CaM-resistant E12 gives lower MyoD binding and higher E12 binding to a MyoD-responsive promoter in vivo and cannot rescue myogenic differentiation that has been inhibited by siRNA against E12 and E47. Our data support the concept that Ca2+-loaded CaM enables myogenesis by inhibiting DNA binding of E-protein homodimers, thereby promoting occupancy of myogenic bHLH protein/E-protein heterodimers on promoters of myogenic target genes.

scholarly journals The basic domain of myogenic basic helix-loop-helix (bHLH) proteins is the novel target for direct inhibition by another bHLH protein, Twist.

1997 ◽  
Vol 17 (11) ◽  
pp. 6563-6573 ◽  
Author(s):  
Y Hamamori ◽  
H Y Wu ◽  
V Sartorelli ◽  
L Kedes
Keyword(s):  
Bhlh Protein ◽  
E Proteins ◽  
Basic Domain ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Protein ◽  
Arginine Residues ◽  
Bhlh Proteins

In vertebrates, the basic helix-loop-helix (bHLH) protein Twist may be involved in the negative regulation of cellular determination and in the differentiation of several lineages, including myogenesis, osteogenesis, and neurogenesis. Although it has been shown that mouse twist (M-Twist) (i) sequesters E proteins, thus preventing formation of myogenic E protein-MyoD complexes and (ii) inhibits the MEF2 transcription factor, a cofactor of myogenic bHLH proteins, overexpression of E proteins and MEF2 failed to rescue the inhibitory effects of M-Twist on MyoD. We report here that M-Twist physically interacts with the myogenic bHLH proteins in vitro and in vivo and that this interaction is required for the inhibition of MyoD by M-Twist. In contrast to the conventional HLH-HLH domain interaction formed in the MyoD/E12 heterodimer, this novel type of interaction uses the basic domains of the two proteins. While the MyoD HLH domain without the basic domain failed to interact with M-Twist, a MyoD peptide containing only the basic and helix 1 regions was sufficient to interact with M-Twist, suggesting that the basic domain contacts M-Twist. The replacement of three arginine residues by alanines in the M-Twist basic domain was sufficient to abolish both the binding and inhibition of MyoD by M-Twist, while the domain retained other M-Twist functions such as heterodimerization with an E protein and inhibition of MEF2 transactivation. These findings demonstrate that M-Twist interacts with MyoD through the basic domains, thereby inhibiting MyoD.

scholarly journals Determinants of Myogenic Specificity within MyoD Are Required for Noncanonical E Box Binding

2007 ◽  
Vol 27 (16) ◽  
pp. 5910-5920 ◽  
Author(s):  
Analeah B. Heidt ◽  
Anabel Rojas ◽  
Ian S. Harris ◽  
Brian L. Black
Keyword(s):  
Transcription Factors ◽  
Promoter Regions ◽  
Basic Domain ◽  
Helix Loop Helix ◽  
E Box ◽  
Muscle Genes ◽  
E Boxes ◽  
Bhlh Transcription Factors

ABSTRACT The MyoD family of basic helix-loop-helix (bHLH) transcription factors has the remarkable ability to induce myogenesis in vitro and in vivo. This myogenic specificity has been mapped to two amino acids in the basic domain, an alanine and threonine, referred to as the myogenic code. These essential determinants of myogenic specificity are conserved in all MyoD family members from worms to humans, yet their function in myogenesis is unclear. Induction of the muscle transcriptional program requires that MyoD be able to locate and stably bind to sequences present in the promoter regions of critical muscle genes. Recent studies have shown that MyoD binds to noncanonical E boxes in the myogenin gene, a critical locus required for myogenesis, through interactions with resident heterodimers of the HOX-TALE transcription factors Pbx1A and Meis1. In the present study, we show that the myogenic code is required for MyoD to bind to noncanonical E boxes in the myogenin promoter and for the formation of a tetrameric complex with Pbx/Meis. We also show that these essential determinants of myogenesis are sufficient to confer noncanonical E box binding to the E12 basic domain. Thus, these data show that noncanonical E box binding correlates with myogenic potential, and we speculate that the myogenic code residues in MyoD function as myogenic determinants via their role in noncanonical E box binding and recognition.

scholarly journals Cloning, purification and preliminary X-ray data analysis of the human ID2 homodimer

2012 ◽  
Vol 68 (11) ◽  
pp. 1354-1358 ◽  
Author(s):  
Marie V. Wong ◽  
Paaventhan Palasingam ◽  
Prasanna R. Kolatkar
Keyword(s):  
Dna Binding ◽  
Potassium Acetate ◽  
Class Iii ◽  
Data Set ◽  
Cell Parameters ◽  
Helix Loop Helix ◽  
Id Protein ◽  
Bhlh Transcription Factors

The ID proteins are named for their role as inhibitors of DNA binding and differentiation. They contain a helix–loop–helix (HLH) domain but lack a basic DNA-binding domain. In complex with basic HLH (bHLH) transcription factors, gene expression is regulated by DNA-binding inactivation. Although the HLH domain is highly conserved and shares a similar topology, the IDs preferentially bind class I bHLH-group members such as E47 (TCF3) but not the class III bHLH member Myc. A structure of an ID protein could potentially shed light on its mechanism. Owing to their short half-livesin vivoand reportedin vitroinstability, this paper describes the strategies that went into expressing sufficient soluble and stable ID2 to finally obtain diffraction-quality crystals. A 2.1 Å resolution data set was collected from a crystal belonging to space groupP3121 with unit-cell parametersa=b= 51.622,c= 111.474 Å, α = β = 90, γ = 120° that was obtained by hanging-drop vapour diffusion in a precipitant solution consisting of 0.1 MMES pH 6.5, 2.0 Mpotassium acetate. The solvent content was consistent with the presence of one or two molecules in the asymmetric unit.

The Basic Helix-Loop-Helix TAL1/SCL and Its Partners E47 and LMO2 Act Upstream of the VE-Cadherin Gene in Endothelial Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1795-1795
Author(s):  
Virginie Deleuze ◽  
Elias Chalhoub ◽  
Rawan El-Hajj ◽  
Christiane Dohet ◽  
Mikael Le Clech ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Ectopic Expression ◽  
Cell Junctions ◽  
Small Interfering Rnas ◽  
Hek 293 ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Cell Cell

Abstract The basic helix-loop-helix protein TAL-1/SCL, essential for the formation of the hematopoietic system, is also required for vascular development and more particularly for embryonic angiogenesis. We previously reported that TAL-1 acts as a positive factor for post-natal angiogenesis by stimulating endothelial morphogenesis. To understand how TAL-1 modulates angiogenesis, we investigated the functional consequences of TAL-1 silencing, mediated by small-interfering RNAs, in human primary endothelial cells (ECs). We found that TAL-1 knockdown impaired in vitro EC tubulomorphogenesis (in 2-D on Matrigel or 3-D in collagen I gel), with the notable absence of cell-cell contacts, a prerequisite for morphogenesis initiation. This cellular deficiency was associated with a dramatic reduction in the vascular-endothelial (VE)-cadherin at intercellular junctions, the major component of endothelial adherens junctions. In contrast, PECAM (or CD31) was present at cell-cell junctions at the same levels as control cells. Importantly, silencing of two known TAL-1-partners in hematopoietic cells, E47 or LMO2, produce the same effects as TAL-1. Accordingly, silencing of TAL-1, as well as E47 and LMO2, provoked down-regulation of VE-cadherin at both the mRNA and protein levels. Transient transfection experiments in HUVECs showed that TAL-1 and E47 regulate the VE-cadherin promoter through a specialized E-box element. Finally, endogenous VE-cadherin transcription could be directly activated in non-endothelial HEK-293 cells that neither express TAL-1 or LMO2, by the sole concomitant ectopic expression of TAL-1, E47 and LMO2. Overall, our data demonstrate that a multiprotein complex containing at least TAL-1, LMO2 and E47 act upstream of the VE-cadherin gene. We are currently performing chromatin immunoprecipitation (ChIP) to investigate whether the TAL-1-containing complex binds in vivo the VE-cadherin promoter. This study identifies VE-cadherin as an upstream TAL-1-target gene in the endothelial lineage, and provides a first clue in TAL-1 function in the control of angiogenesis.

Identification of Key Cellular Regulators That Interact with Id1 To Control Hematopoietic Stem Cell Behavior.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1319-1319
Author(s):  
Vladimir Jankovic ◽  
Alessia Ciarrocchi ◽  
Tony DeBlasio ◽  
Robert Benezra ◽  
Stephen D. Nimer
Keyword(s):  
Transcription Factors ◽  
Transcriptional Repression ◽  
Genetic Interaction ◽  
Dominant Negative ◽  
Double Knockout ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Helix Loop Helix

Abstract The ability of hematopoietic stem cells to tightly regulate the transition from relative quiescence and self-renewal to the transiently amplifying, differentiating progenitor fate is critical for HSC homeostasis as well as their regenerative capacity. We have recently described the diminished frequency and rapid exhaustion of HSC self-renewal capacity in the absence of the dominant negative helix-loop-helix molecule Id1. Furthermore, Id1 null HSCs have an increased rate of cycling, coupled with accelerated myeloid commitment both in vivo and in vitro. This is reflected in the elevated expression of myelo-erythroid transcription factors (c/EBPalpha and GATA1) within the Lin−c-kit+Sca-1+ population - “myeloid priming”. The major targets of Id1 mediated transcriptional repression are the ubiquitous E protein E2A as well as Ets transcription factors (Ets1 and Ets2). We hypothesized that the unrestrained activity of these and/or other targets of Id1 transcriptional repression leads to premature HSC commitment in Id1 null animals. Indeed, we show that HSC differentiation in culture can be delayed by transduction of E2A directed shRNA specifically in Id1 null, but not in wild-type Id1 expressing cells. This indicates an abnormal E2A activity in Id1 null HSCs that could be responsible for their increased differentiation status. To further define the transcriptional deregulation in Id1 null HSCs, we have used the Affymetrix microarray technology. We observed ~3 fold increased expression of the CDK inhibitor p21 in freshly isolated Id1 null HSCs and have confirmed this result by multiple independent qPCR measurements. The transcriptional induction of p21 by E2A as well as its repression by Id1 have been well established. Therefore, the observed p21 induction could be explained by the elevated level of E2A activity in HSCs in the absence of Id1 expression. To explore the functional significance of Id1 mediated p21 regulation in HSCs, we have generated p21/Id1 double knockout animals. Surprisingly, despite its reported function in restricting the cell cycle entry of normal HSCs, we show that in the context of Id1 loss, p21 expression is required for the accelerated HSC cycling, and unlike Id1 single null HSCs, p21/Id1 double knockout HSCs do not show accelerated myeloid differentiation in culture. Therefore, we propose that Id1 actively represses E2A activity in HSCs, as well as the induction of p21, which could be an important component of the HSC commitment program. Further studies will be presented defining the in vivo relevance of the Id1/p21 genetic interaction for HSC growth and differentiation.

Myostatin silencing effect on basic helix–loop–helix transcription factors in caprine foetal fibroblast cells

2017 ◽  
Author(s):  
Biswajyoti Borah ◽  
Ajit Pratap Singh ◽  
Hamen Gogoi ◽  
Amlan Jyoti Phukan ◽  
Bikash Chandra Sarkhel
Keyword(s):  
Transcription Factors ◽  
Muscle Development ◽  
Pearson Correlation ◽  
Negative Regulator ◽  
Fibroblast Cells ◽  
Animal Cells ◽  
Myostatin Gene ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix

Transgenic food animal production is one of the potential and need oriented research to mitigate the food crises of the world. In vitro gene silenced animal cells and making use of these cells for transgenesis one of the suitable way to produce productive animals. Myostatin is a negative regulator of muscle growth, has the potential to increase the muscle mass upon its silencing. Four Hush 29-mer anti- myostatin (MSTN) shRNA constructs were checked for myostatin gene silencing in caprine foetal fibroblast cells and its subsequent effect on basic helix– loop–helix (bHLH) transcription factors. These factors are necessary for the terminal differentiation, proliferation, and homeostasis of muscle development. Different shRNA constructs displayed 55.1 to 91.5% (p less than 0.01) of myostatin silencing in caprine foetal fibroblast cells and upregulation of myogenic gene. Upregulation of 7.97 to 111.67 % for MyoD, 77.0 % to 319.47 % for myogenin, 16.67 % to 138.0 % for Myf5 were observed . The Pearson correlation established a negative correlation between myostatin and genes under study. Result suggests that knockdown of MSTN a potential approach to improve caprine musculatures.

Sign in / Sign up

Export Citation Format

Share Document