scholarly journals Atonal homolog 7 (ATOH7) loss-of-function mutations in predominant bilateral optic nerve hypoplasia

2019 ◽  
Author(s):  
David Grubich Atac ◽  
Samuel Koller ◽  
James V M Hanson ◽  
Silke Feil ◽  
Amit Tiwari ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Optic Nerve ◽  
Transcriptional Activation ◽  
Functional Characterization ◽  
Optic Nerve Hypoplasia ◽  
Luciferase Reporter ◽  
Bhlh Transcription Factor ◽  
Compound Heterozygous ◽  
Missense Mutations ◽  
Single Nucleotide Variants

Abstract Optic nerve hypoplasia (ONH) is a congenital optic nerve abnormality caused by underdevelopment of retinal ganglion cells (RGCs). Despite being a rare disease, ONH is the most common optic disc anomaly in ophthalmological practice. So far, mutations in several genes have been identified as causative, however many cases of ONH remain without a molecular explanation. The early transcription factor atonal basic-helix-loop-helix (bHLH) transcription factor 7 (ATOH7) is expressed in retinal progenitor cells and has a crucial role in RGC development. Previous studies have identified several mutations in the ATOH7 locus in cases of eye developmental diseases such as nonsyndromic congenital retinal nonattachment and persistent hyperplasia of the primary vitreous. Here we present two siblings with a phenotype predominated by bilateral ONH, with additional features of foveal hypoplasia and distinct vascular abnormalities, where whole-exome sequencing identified two compound heterozygous missense mutations affecting a conserved amino acid residue within the bHLH domain of ATOH7 (NM_145178.3:c.175G>A; p.(Ala59Thr) and c.176C>T; p.(Ala59Val)). ATOH7 expression constructs with patient single nucleotide variants were cloned for functional characterization. Protein analyses revealed decreased protein amounts and significantly enhanced degradation in the presence of E47, a putative bHLH dimerization partner. Protein interaction assays revealed decreased heterodimerization and DNA-binding of ATOH7 variants, resulting in total loss of transcriptional activation of luciferase reporter gene expression. These findings strongly support pathogenicity of the two ATOH7 mutations, one of which is novel. Additionally, this report highlights the possible impact of altered ATOH7 dimerization on protein stability and function.

scholarly journals The bHLH transcription factor BIS1 controls the iridoid branch of the monoterpenoid indole alkaloid pathway in Catharanthus roseus

2015 ◽  
Vol 112 (26) ◽  
pp. 8130-8135 ◽  
Author(s):  
Alex Van Moerkercke ◽  
Priscille Steensma ◽  
Fabian Schweizer ◽  
Jacob Pollier ◽  
Ivo Gariboldi ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Catharanthus Roseus ◽  
Transcriptional Activation ◽  
Indole Alkaloid ◽  
Bioactive Metabolites ◽  
Bhlh Transcription Factor ◽  
Ethylene Response Factor ◽  
Madagascar Periwinkle ◽  
Genes Encoding ◽  
Monoterpenoid Indole Alkaloid

Plants make specialized bioactive metabolites to defend themselves against attackers. The conserved control mechanisms are based on transcriptional activation of the respective plant species-specific biosynthetic pathways by the phytohormone jasmonate. Knowledge of the transcription factors involved, particularly in terpenoid biosynthesis, remains fragmentary. By transcriptome analysis and functional screens in the medicinal plant Catharanthus roseus (Madagascar periwinkle), the unique source of the monoterpenoid indole alkaloid (MIA)-type anticancer drugs vincristine and vinblastine, we identified a jasmonate-regulated basic helix–loop–helix (bHLH) transcription factor from clade IVa inducing the monoterpenoid branch of the MIA pathway. The bHLH iridoid synthesis 1 (BIS1) transcription factor transactivated the expression of all of the genes encoding the enzymes that catalyze the sequential conversion of the ubiquitous terpenoid precursor geranyl diphosphate to the iridoid loganic acid. BIS1 acted in a complementary manner to the previously characterized ethylene response factor Octadecanoid derivative-Responsive Catharanthus APETALA2-domain 3 (ORCA3) that transactivates the expression of several genes encoding the enzymes catalyzing the conversion of loganic acid to the downstream MIAs. In contrast to ORCA3, overexpression of BIS1 was sufficient to boost production of high-value iridoids and MIAs in C. roseus suspension cell cultures. Hence, BIS1 might be a metabolic engineering tool to produce sustainably high-value MIAs in C. roseus plants or cultures.

scholarly journals Diversification of Transcription Factor NF-κB in Protists

2021 ◽  
Author(s):  
Leah M. Williams ◽  
Sainetra Sridhar ◽  
Jason Samaroo ◽  
Ebubechi K. Adindu ◽  
Anvitha Addanki ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Functional Characterization ◽  
Binding Activity ◽  
Life Stages ◽  
Rna Seq ◽  
Dna Binding Activity ◽  
Inhibitory Domain

In this report, we investigate the evolution of transcription factor NF-κB by examining its structure, activity, and regulation in two protists using phylogenetic, cellular, and biochemical techniques. In Capsaspora owczarzaki (Co), we find that full-length NF-κB has an N-terminal DNA-binding domain and a C-terminal Ankyrin (ANK) repeat inhibitory domain, and its DNA-binding activity is more similar to metazoan NF-κB rather than Rel proteins. As with mammalian NF-κB proteins, removal of the ANK repeats is required for Co-NF-κB to enter the nucleus, bind DNA, and activate transcription. However, C-terminal processing of Co-NF-κB is not induced by co-expression of IKK in human cells. Exogenously expressed Co-NF-κB localizes to the nucleus in Co cells. NF-κB mRNA and DNA-binding levels differ across three life stages of Capsaspora, suggesting distinct roles for NF-κB in these life stages. RNA-seq and GO analyses identify possible gene targets and biological functions of Co-NF-κB. We also show that three NF-κB-like proteins from the choanoflagellate Acanthoeca spectabilis (As) all consist of primarily the N-terminal conserved Rel Homology domain sequences of NF-κB, and lack C-terminal ANK repeats. All three As-NF-κB proteins constitutively enter the nucleus of human and Co cells, but differ in their DNA-binding and transcriptional activation activities. Furthermore, all three As-NF-κB proteins can form heterodimers, indicating that NF-κB diversified into multi-subunit families at least two times during evolution. Overall, these results present the first functional characterization of NF-κB in a taxonomic kingdom other than Animalia and provide information about the evolution and diversification of this biologically important transcription factor.

scholarly journals The E2F transcription factor activates a replication-dependent human H2A gene in early S phase of the cell cycle.

1996 ◽  
Vol 16 (5) ◽  
pp. 1889-1895 ◽  
Author(s):  
F Oswald ◽  
T Dobner ◽  
M Lipp
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Transcription Factor ◽  
Transcriptional Activation ◽  
Promoter Region ◽  
S Phase ◽  
Histone Gene ◽  
Luciferase Reporter ◽  
Transcriptional Level ◽  
Histone Gene Expression

Histone gene expression is restricted to the S phase of the cell cycle. Control is mediated by a complex network of sequence-specific DNA-binding factors and protein-protein interactions in response to cell cycle progression. To further investigate the regulatory functions that are associated at the transcriptional level, we analyzed the regulation of a replication-dependent human H2A.1-H2B.2 gene pair. We found that transcription factor E2F binds specifically to an E2F recognition motif in the H2A.1 promoter region. Activation of the H2A.1 promoter by E2F-1 was shown by use of luciferase reporter constructs of the intergenic promoter region. Overexpression of the human retinoblastoma suppressor gene product RB suppressed E2F-1 mediated transcriptional activation, indicating an E2F-dependent regulation of promoter activity during the G1-to-S-phase transition. Furthermore, the activity of the H2A.1 promoter was also downregulated by overexpression of the RB-related p107, a protein that has been detected in S-phase-specific protein complexes of cyclin A, E2F, and cdk2. In synchronized HeLa cells, expression of luciferase activity was induced at the beginning of DNA synthesis and was dependent on the presence of an E2F-binding site in the H2A.1 promoter. Together with the finding that E2F-binding motifs are highly conserved in H2A promoters of other species, our results suggest that E2F plays an important role in the coordinate regulation of S-phase-specific histone gene expression.

Alternative splicing yields novel BMAL2 variants: tissue distribution and functional characterization

2002 ◽  
Vol 283 (1) ◽  
pp. C103-C114 ◽  
Author(s):  
John A. Schoenhard ◽  
Mesut Eren ◽  
Carl H. Johnson ◽  
Douglas E. Vaughan
Keyword(s):  
Alternative Splicing ◽  
Transcriptional Activation ◽  
Functional Characterization ◽  
Plasminogen Activator Inhibitor ◽  
Luciferase Reporter ◽  
Plasminogen Activator Inhibitor 1 ◽  
Luciferase Reporter Gene ◽  
Helix Loop Helix ◽  
Peripheral Oscillators ◽  
Physiological Processes

The BMAL2 gene encodes a member of the basic helix-loop-helix PER-ARNT-SIM family of transcription factors, which control diverse physiological processes including circadian rhythms. We identified four novel human BMAL2 transcripts that differ by alternative splicing within their NH2-terminal regions. Divergent expression of these and previously reported transcripts was observed among human tissues. The functional consequences of alternative splicing for transcriptional activation by CLOCK:BMAL2 heterodimers were assessed using luciferase reporter gene constructs that contained one of three diurnally regulated promoters, namely, those of the mouse period1, mouse vasopressin, and human plasminogen activator inhibitor-1 genes. These studies revealed that alternative splicing generates BMAL2 isoforms possessing high, medium, low, or no transcriptional activity. Similar results were obtained with each promoter, suggesting that alternative splicing may influence the amplitudes of both central and peripheral oscillators. Indeed, alternative splicing of BMAL2 may provide tissues with a rheostat capable of regulating CLOCK:BMAL2 heterodimer function across a broad continuum of potential transcriptional activities to accommodate varied metabolic demands and physiological roles.

The molecular cloning and functional characterization of MdMYC2, a bHLH transcription factor in apple

2016 ◽  
Vol 108 ◽  
pp. 24-31 ◽  
Author(s):  
Jian-Ping An ◽  
Hao-Hao Li ◽  
Lai-Qing Song ◽  
Ling Su ◽  
Xin Liu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Molecular Cloning ◽  
Functional Characterization ◽  
Bhlh Transcription Factor

scholarly journals Comparison of NF-κB from the protists Capsaspora owczarzaki and Acanthoeca spectabilis reveals extensive evolutionary diversification of this transcription factor

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Leah M. Williams ◽  
Sainetra Sridhar ◽  
Jason Samaroo ◽  
Jada Peart ◽  
Ebubechi K. Adindu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Functional Characterization ◽  
Evolutionary Diversification ◽  
Evolutionary Origins ◽  
Dna Binding Specificity ◽  
Higher Taxa ◽  
Rel Homology Domain

AbstractWe provide a functional characterization of transcription factor NF-κB in protists and provide information about the evolution and diversification of this biologically important protein. We characterized NF-κB in two protists using phylogenetic, cellular, and biochemical techniques. NF-κB of the holozoan Capsaspora owczarzaki (Co) has an N-terminal DNA-binding domain and a C-terminal Ankyrin repeat (ANK) domain, and its DNA-binding specificity is more similar to metazoan NF-κB proteins than to Rel proteins. Removal of the ANK domain allows Co-NF-κB to enter the nucleus, bind DNA, and activate transcription. However, C-terminal processing of Co-NF-κB is not induced by IκB kinases in human cells. Overexpressed Co-NF-κB localizes to the cytoplasm in Co cells. Co-NF-κB mRNA and DNA-binding levels differ across three Capsaspora life stages. RNA-sequencing and GO analyses identify possible gene targets of Co-NF-κB. Three NF-κB-like proteins from the choanoflagellate Acanthoeca spectabilis (As) contain conserved Rel Homology domain sequences, but lack C-terminal ANK repeats. All three As-NF-κB proteins constitutively enter the nucleus of cells, but differ in their DNA-binding abilities, transcriptional activation activities, and dimerization properties. These results provide a basis for understanding the evolutionary origins of this key transcription factor and could have implications for the origins of regulated immunity in higher taxa.

scholarly journals Functional Characterization of a Sugar Beet BvbHLH93 Transcription Factor in Salt Stress Tolerance

2021 ◽  
Vol 22 (7) ◽  
pp. 3669
Author(s):  
Yuguang Wang ◽  
Shuang Wang ◽  
Ye Tian ◽  
Qiuhong Wang ◽  
Sixue Chen ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Antioxidant Enzymes ◽  
Salt Stress ◽  
Sugar Beet ◽  
Stress Tolerance ◽  
Stress Responses ◽  
Functional Characterization ◽  
Bhlh Transcription Factor ◽  
Salt Stress Tolerance ◽  
Antioxidant Genes

The basic/helix–loop–helix (bHLH) transcription factor (TF) plays an important role for plant growth, development, and stress responses. Previously, proteomics of NaCl treated sugar beet leaves revealed that a bHLH TF, BvbHLH93, was significantly increased under salt stress. The BvbHLH93 protein localized in the nucleus and exhibited activation activity. The expression of BvbHLH93 was significantly up-regulated in roots and leaves by salt stress, and the highest expression level in roots and leaves was 24 and 48 h after salt stress, respectively. Furthermore, constitutive expression of BvbHLH93 conferred enhanced salt tolerance in Arabidopsis, as indicated by longer roots and higher content of chlorophyll than wild type. Additionally, the ectopic expression lines accumulated less Na+ and MDA, but more K+ than the WT. Overexpression of the BvBHLH93 enhanced the activities of antioxidant enzymes by positively regulating the expression of antioxidant genes SOD and POD. Compared to WT, the overexpression plants also had low expression levels of RbohD and RbohF, which are involved in reactive oxygen species (ROS) production. These results suggest that BvbHLH93 plays a key role in enhancing salt stress tolerance by enhancing antioxidant enzymes and decreasing ROS generation.

scholarly journals Mutations in LAMB2 Are Associated With Albuminuria and Optic Nerve Hypoplasia With Hypopituitarism

2019 ◽  
Vol 105 (3) ◽  
pp. 595-599
Author(s):  
Mona Tahoun ◽  
Jennifer C Chandler ◽  
Emma Ashton ◽  
Scott Haston ◽  
Athia Hannan ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Optic Nerve ◽  
Basement Membrane ◽  
Anterior Pituitary ◽  
Congenital Nephrotic Syndrome ◽  
Optic Nerve Hypoplasia ◽  
Hormone Deficiency ◽  
Compound Heterozygous ◽  
Macroscopic Hematuria ◽  
Laminin Β2

Abstract Context Mutations in LAMB2, encoding the basement membrane protein, laminin β2, are associated with an autosomal recessive disorder characterized by congenital nephrotic syndrome, ocular abnormalities, and neurodevelopmental delay (Pierson syndrome). Case description This report describes a 12-year-old boy with short stature, visual impairment, and developmental delay who presented with macroscopic hematuria and albuminuria. He had isolated growth hormone deficiency, optic nerve hypoplasia, and a small anterior pituitary with corpus callosum dysgenesis on his cranial magnetic resonance imaging, thereby supporting a diagnosis of optic nerve hypoplasia syndrome. Renal histopathology revealed focal segmental glomerulosclerosis. Using next-generation sequencing on a targeted gene panel for steroid-resistant nephrotic syndrome, compound heterozygous missense mutations were identified in LAMB2 (c.737G>A p.Arg246Gln, c.3982G>C p.Gly1328Arg). Immunohistochemical analysis revealed reduced glomerular laminin β2 expression compared to control kidney and a thin basement membrane on electron microscopy. Laminin β2 is expressed during pituitary development and Lamb2–/– mice exhibit stunted growth, abnormal neural retinae, and here we show, abnormal parenchyma of the anterior pituitary gland. Conclusion We propose that patients with genetically undefined optic nerve hypoplasia syndrome should be screened for albuminuria and, if present, screened for mutations in LAMB2.

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