scholarly journals Dissection of the cellular function of the ZBED6 transcription factor in mouse myoblast cells using gene editing, RNAseq and proteomics

2019 ◽  
Author(s):  
Shady Younis ◽  
Rakan Naboulsi ◽  
Xuan Wang ◽  
Xiaofang Cao ◽  
Mårten Larsson ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Gene Editing ◽  
Muscle Development ◽  
Myoblast Differentiation ◽  
Mitochondrial Activity ◽  
Ras Signaling ◽  
Altered Expression ◽  
Significant Enrichment ◽  
Myotube Hypertrophy ◽  
Myoblast Cells

SUMMARYThe transcription factor ZBED6 acts as a repressor of Igf2 and affects directly or indirectly the transcriptional regulation of thousands of genes. Here, we use gene editing in mouse C2C12 myoblasts and show that ZBED6 regulates Igf2 exclusively through its binding site 5′-GGCTCG-3′ in intron 1 of Igf2. Deletion of this motif (Igf2ΔGGCT) or complete ablation of Zbed6 leads to ~20-fold up-regulation of IGF2 protein. Quantitative proteomics revealed an activation of Ras signaling pathway in both Zbed6-/- and Igf2ΔGGCT myoblasts, and a significant enrichment of mitochondrial membrane proteins among proteins showing altered expression in Zbed6-/- myoblasts. Both Zbed6-/- and Igf2ΔGGCT myoblasts showed a faster growth rate and developed myotube hypertrophy. These cells exhibited an increased O2 consumption rate, due to IGF2 up-regulation. Transcriptome analysis revealed ~30% overlap between differentially expressed genes in Zbed6-/- and Igf2ΔGGCT myotubes, with an enrichment of up-regulated genes involved in muscle development. In contrast, ZBED6-overexpression in myoblasts led to cell apoptosis, cell cycle arrest, reduced mitochondrial activities and ceased myoblast differentiation. The similarities in growth and differentiation phenotypes observed in Zbed6-/- and Igf2ΔGGCT myoblasts demonstrates that ZBED6 affects mitochondrial activity and myogenesis largely through its regulation of IGF2 expression. This study suggests that the interaction between ZBED6-Igf2 may be a therapeutic target for human diseases where anabolism is impaired.

Abstract 344: Finding the Achilles’ heel of Muscle Giant---TALEN-mediated Gene-editing in Zebrafish Titin

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Yu-Huan Shih ◽  
Xiaolei Xu
Keyword(s):  
Alternative Splicing ◽  
Gene Editing ◽  
Muscle Development ◽  
Exon Skipping ◽  
Transcription Activator ◽  
Detailed Characterization ◽  
A Domains ◽  
Underlying Mechanisms ◽  
Sarcomere Assembly

Background: TITIN (TTN) has more than 300 exons and encodes a gigantic protein that is crucial for heart and muscle development. Mutations in TTN caused a variety of human diseases including cardiomyopathy and muscular dystrophy. Recently, dilated cardiomyopathy-associated mutations on TTN have been found more frequently in exons encoding A-band domains but less in exons encoding the N-terminal Z-disc domains, suggesting that mutations in different exons of TTN cause distinct consequences. To elucidate the underlying mechanisms, we leveraged the Transcription Activator-Like Effects Nuclease (TALEN) technology in zebrafish to introduce truncating mutations in different exons of ttn, and then study their effects on heart and somites. Results: We generated truncational mutations in different exons of zebrafish titins encoding Z-disc, N2B, Novex-3, and A domains, respectively. Because zebrafish contains two titin homologues, ttna and ttnb, we introduced mutations in both genes at the corresponding loci. While both Z-disc and A band mutations on ttna disrupted sarcomere assembly in heart and somites, Z-disc or A band mutations on ttnb only affect somites without affecting the heart. Interestingly, a Z-disc mutation on ttna resulted in milder phenotypes than an A-band mutation, while a Z-disc mutation on ttnb generated severer phenotypes than an A-band mutation. No phenotype was observed in the homozygous fish in either ttna-novex-3 or ttnb-N2B mutant fish. Conclusions: A spectrum of truncational mutations in ttna and ttnb has been generated in zebrafish using the TALEN technology. Mutations in different exons result in different phenotypes. Detailed characterization of these mutants and double mutants will be presented, which shall elicit distinct contribution of alternative splicing and exon skipping as two candidate mechanisms during pathogenesis of Titinopathies.

Isolation and characterization of a downstream target of Pax6 in the mammalian retinal primordium

Development ◽  
2001 ◽  
Vol 128 (20) ◽  
pp. 3987-3994 ◽  
Author(s):  
Gilbert Bernier ◽  
Wolfgang Vukovich ◽  
Lorenz Neidhardt ◽  
Bernhard G. Herrmann ◽  
Peter Gruss
Keyword(s):  
Transcription Factor ◽  
Expression Pattern ◽  
Large Scale ◽  
Ectopic Expression ◽  
Signal Transduction Pathway ◽  
Homeobox Gene ◽  
Optic Vesicle ◽  
Altered Expression ◽  
Retina Development ◽  
Isolation And Characterization

The transcription factor Pax6 is required for eye morphogenesis in humans, mice and insects, and can induce ectopic eye formation in vertebrate and invertebrate organisms. Although the role of Pax6 has intensively been studied, only a limited number of genes have been identified that depend on Pax6 activity for their expression in the mammalian visual system. Using a large-scale in situ hybridization screen approach, we have identified a novel gene expressed in the mouse optic vesicle. This gene, Necab, encodes a putative cytoplasmic Ca2+-binding protein and coincides with Pax6 expression pattern in the neural ectoderm of the optic vesicle and in the forebrain pretectum. Remarkably, Necab expression is absent in both structures in Pax6 mutant embryos. By contrast, the optic vesicle-expressed homeobox genes Rx, Six3, Otx2 and Lhx2 do not exhibit an altered expression pattern. Using gain-of-function experiments, we show that Pax6 can induce ectopic expression of Necab, suggesting that Necab is a direct or indirect transcriptional target of Pax6. In addition, we have found that Necab misexpression can induce ectopic expression of the homeobox gene Chx10, a transcription factor implicated in retina development. Taken together, our results provide evidence that Necab is genetically downstream of Pax6 and that it is a part of a signal transduction pathway in retina development.

Altered expression of the bZIP transcription factor DRINK ME affects growth and reproductive development inArabidopsis thaliana

2016 ◽  
Vol 88 (3) ◽  
pp. 437-451 ◽  
Author(s):  
Paulina Lozano-Sotomayor ◽  
Ricardo A. Chávez Montes ◽  
Marina Silvestre-Vañó ◽  
Humberto Herrera-Ubaldo ◽  
Raffaella Greco ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Reproductive Development ◽  
Bzip Transcription Factor ◽  
Altered Expression

scholarly journals Genome-Based Identification of Cancer Genes by Proviral Tagging in Mouse Retrovirus-Induced T-Cell Lymphomas

2003 ◽  
Vol 77 (3) ◽  
pp. 2056-2062 ◽  
Author(s):  
Rachel Kim ◽  
Alla Trubetskoy ◽  
Takeshi Suzuki ◽  
Nancy A. Jenkins ◽  
Neal G. Copeland ◽  
...  
Keyword(s):  
T Cell ◽  
Molecular Mechanisms ◽  
Mouse Genome ◽  
Human Cancer ◽  
Inverse Pcr ◽  
Ras Signaling ◽  
Cancer Genes ◽  
Altered Expression ◽  
New Genes ◽  
T Cell Lymphomas

ABSTRACT The identification of tumor-inducing genes is a driving force for elucidating the molecular mechanisms underlying cancer. Many retroviruses induce tumors by insertion of viral DNA adjacent to cellular oncogenes, resulting in altered expression and/or structure of the encoded proteins. The availability of the mouse genome sequence now allows analysis of retroviral common integration sites in murine tumors to be used as a genetic screen for identification of large numbers of candidate cancer genes. By positioning the sequences of inverse PCR-amplified, virus-host junction fragments within the mouse genome, 19 target genes were identified in T-cell lymphomas induced by the retrovirus SL3-3. The candidate cancer genes included transcription factors (Fos, Gfi1, Lef1, Myb, Myc, Runx3, and Sox3), all three D cyclins, Ras signaling pathway components (Rras2/TC21 and Rasgrp1), and Cmkbr7/CCR7. The most frequent target was Rras2. Insertions as far as 57 kb away from the transcribed portion were associated with substantially increased transcription of Rras2, and no coding sequence mutations, including those typically involved in Ras activation, were detected. These studies demonstrate the power of genome-based analysis of retroviral insertion sites for cancer gene discovery, identify several new genes worth examining for a role in human cancer, and implicate the pathways in which those genes act in lymphomagenesis. They also provide strong genetic evidence that overexpression of unmutated Rras2 contributes to tumorigenesis, thus suggesting that it may also do so if it is inappropriately expressed in human tumors.

Overlapping functions of the myogenic bHLH genes MRF4 and MyoD revealed in double mutant mice

Development ◽  
1998 ◽  
Vol 125 (13) ◽  
pp. 2349-2358 ◽  
Author(s):  
A. Rawls ◽  
M.R. Valdez ◽  
W. Zhang ◽  
J. Richardson ◽  
W.H. Klein ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Double Mutant ◽  
Muscle Development ◽  
Expression Patterns ◽  
Muscle Fibers ◽  
Mutant Mice ◽  
Myoblast Differentiation ◽  
Bhlh Genes ◽  
Bhlh Factors

The myogenic basic helix-loop-helix (bHLH) genes - MyoD, Myf5, myogenin and MRF4 - exhibit distinct, but overlapping expression patterns during development of the skeletal muscle lineage and loss-of-function mutations in these genes result in different effects on muscle development. MyoD and Myf5 have been shown to act early in the myogenic lineage to establish myoblast identity, whereas myogenin acts later to control myoblast differentiation. In mice lacking myogenin, there is a severe deficiency of skeletal muscle, but some residual muscle fibers are present in mutant mice at birth. Mice lacking MRF4 are viable and have skeletal muscle, but they upregulate myogenin expression, which could potentially compensate for the absence of MRF4. Previous studies in which Myf5 and MRF4 null mutations were combined suggested that these genes do not share overlapping myogenic functions in vivo. To determine whether the functions of MRF4 might overlap with those of myogenin or MyoD, we generated double mutant mice lacking MRF4 and either myogenin or MyoD. MRF4/myogenin double mutant mice contained a comparable number of residual muscle fibers to mice lacking myogenin alone and myoblasts from those double mutant mice formed differentiated multinucleated myotubes in vitro as efficiently as wild-type myoblasts, indicating that neither myogenin nor MRF4 is absolutely essential for myoblast differentiation. Whereas mice lacking either MRF4 or MyoD were viable and did not show defects in muscle development, MRF4/MyoD double mutants displayed a severe muscle deficiency similar to that in myogenin mutants. Myogenin was expressed in MRF4/MyoD double mutants, indicating that myogenin is insufficient to support normal myogenesis in vivo. These results reveal unanticipated compensatory roles for MRF4 and MyoD in the muscle differentiation pathway and suggest that a threshold level of myogenic bHLH factors is required to activate muscle structural genes, with this level normally being achieved by combinations of multiple myogenic bHLH factors.

scholarly journals Chlorella vulgaris Improves the Regenerative Capacity of Young and Senescent Myoblasts and Promotes Muscle Regeneration

2019 ◽  
Vol 2019 ◽  
pp. 1-16 ◽  
Author(s):  
Nurhazirah Zainul Azlan ◽  
Yasmin Anum Mohd Yusof ◽  
Ekram Alias ◽  
Suzana Makpol
Keyword(s):  
Chlorella Vulgaris ◽  
Muscle Regeneration ◽  
Myogenic Differentiation ◽  
Myoblast Differentiation ◽  
Regenerative Capacity ◽  
Maturation Index ◽  
Myoblast Cells ◽  
And Function ◽  
Pharmaceutical Uses

Sarcopenia is characterized by the loss of muscle mass, strength, and function with ageing. With increasing life expectancy, greater attention has been given to counteracting the effects of sarcopenia on the growing elderly population. Chlorella vulgaris, a microscopic, unicellular, green alga with the potential for various pharmaceutical uses, has been widely studied in this context. This study is aimed at determining the effects of C. vulgaris on promoting muscle regeneration by evaluating myoblast regenerative capacity in vitro. Human skeletal myoblast cells were cultured and underwent serial passaging into young and senescent phases and were then treated with C. vulgaris, followed by the induction of differentiation. The ability of C. vulgaris to promote myoblast differentiation was analysed through cellular morphology, real-time monitoring, cell proliferation, senescence-associated β-galactosidase (SA-β-gal) expression, myogenic differentiation, myogenin expression, and cell cycle profiling. The results obtained showed that senescent myoblasts exhibited an enlarged and flattened morphology, with increased SA-β-gal expression, reduced myogenic differentiation, decreased expression of myogenin, and an increased percentage of cells in the G0/G1 phase. Treatment with C. vulgaris resulted in decreased SA-β-gal expression and promotion of myogenic differentiation, as observed via an increased fusion index, maturation index, myotube size, and surface area and an increased percentage of cells that stained positive for myogenin. In conclusion, C. vulgaris improves the regenerative capacity of young and senescent myoblasts and promotes myoblast differentiation, indicating its potential to promote muscle regeneration.

scholarly journals Myoferlin Regulates Wnt/β-Catenin Signaling-Mediated Skeletal Muscle Development by Stabilizing Dishevelled-2 Against Autophagy

2019 ◽  
Vol 20 (20) ◽  
pp. 5130 ◽  
Author(s):  
Shunshun Han ◽  
Can Cui ◽  
Haorong He ◽  
Xiaoxu Shen ◽  
Yuqi Chen ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Atrophy ◽  
Muscle Development ◽  
Skeletal Muscle Atrophy ◽  
Myoblast Differentiation ◽  
C2c12 Myoblast ◽  
Skeletal Muscle Development ◽  
Muscle Tissues ◽  
Underlying Mechanisms ◽  
Canonical Wnt

Myoferlin (MyoF), which is a calcium/phospholipid-binding protein expressed in cardiac and muscle tissues, belongs to the ferlin family. While MyoF promotes myoblast differentiation, the underlying mechanisms remain poorly understood. Here, we found that MyoF not only promotes C2C12 myoblast differentiation, but also inhibits muscle atrophy and autophagy. In the present study, we found that myoblasts fail to develop into mature myotubes due to defective differentiation in the absence of MyoF. Meanwhile, MyoF regulates the expression of atrophy-related genes (Atrogin-1 and MuRF1) to rescue muscle atrophy. Furthermore, MyoF interacts with Dishevelled-2 (Dvl-2) to activate canonical Wnt signaling. MyoF facilitates Dvl-2 ubiquitination resistance by reducing LC3-labeled Dvl-2 levels and antagonizing the autophagy system. In conclusion, we found that MyoF plays an important role in myoblast differentiation during skeletal muscle atrophy. At the molecular level, MyoF protects Dvl-2 against autophagy-mediated degradation, thus promoting activation of the Wnt/β-catenin signaling pathway. Together, our findings suggest that MyoF, through stabilizing Dvl-2 and preventing autophagy, regulates Wnt/β-catenin signaling-mediated skeletal muscle development.

scholarly journals What turns CREB on? And off? And why does it matter?

2020 ◽  
Vol 77 (20) ◽  
pp. 4049-4067 ◽  
Author(s):  
André Steven ◽  
Michael Friedrich ◽  
Paul Jank ◽  
Nadine Heimer ◽  
Jan Budczies ◽  
...  
Keyword(s):  
Overall Survival ◽  
Transcription Factor ◽  
Cyclic Amp ◽  
Therapeutic Target ◽  
Therapy Response ◽  
Predictive Marker ◽  
Tumor Patients ◽  
Altered Expression ◽  
Underlying Mechanisms ◽  
And Function

Abstract Altered expression and function of the transcription factor cyclic AMP response-binding protein (CREB) has been identified to play an important role in cancer and is associated with the overall survival and therapy response of tumor patients. This review focuses on the expression and activation of CREB under physiologic conditions and in tumors of distinct origin as well as the underlying mechanisms of CREB regulation by diverse stimuli and inhibitors. In addition, the clinical relevance of CREB is summarized, including its use as a prognostic and/or predictive marker as well as a therapeutic target.

scholarly journals Transcription Factor CTIP2 Maintains Hair Follicle Stem Cell Pool and Contributes to Altered Expression of LHX2 and NFATC1

2015 ◽  
Vol 135 (11) ◽  
pp. 2593-2602 ◽  
Author(s):  
Shreya Bhattacharya ◽  
Heather Wheeler ◽  
Mark Leid ◽  
Gitali Ganguli-Indra ◽  
Arup K. Indra
Keyword(s):  
Transcription Factor ◽  
Stem Cell ◽  
Hair Follicle ◽  
Altered Expression ◽  
Cell Pool ◽  
Hair Follicle Stem Cell ◽  
Stem Cell Pool

scholarly journals Zfp422 promotes skeletal muscle differentiation by regulating EphA7 to induce appropriate myoblast apoptosis

2019 ◽  
Vol 27 (5) ◽  
pp. 1644-1659 ◽  
Author(s):  
Yaping Nie ◽  
Shufang Cai ◽  
Renqiang Yuan ◽  
Suying Ding ◽  
Xumeng Zhang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Zinc Finger ◽  
Muscle Development ◽  
Zinc Finger Protein ◽  
Critical Role ◽  
Muscle Differentiation ◽  
C2c12 Cells ◽  
Myoblast Differentiation ◽  
Skeletal Muscle Differentiation ◽  
Finger Protein

Abstract Zinc finger protein 422 (Zfp422) is a widely expressed zinc finger protein that serves as a transcriptional factor to regulate downstream gene expression, but until now, little is known about its roles in myogenesis. We found here that Zfp422 plays a critical role in skeletal muscle development and regeneration. It highly expresses in mouse skeletal muscle during embryonic development. Specific knockout of Zfp422 in skeletal muscle impaired embryonic muscle formation. Satellite cell-specific Zfp422 deletion severely inhibited muscle regeneration. Myoblast differentiation and myotube formation were suppressed in Zfp422-deleted C2C12 cells, isolated primary myoblasts, and satellite cells. Chromatin Immunoprecipitation Sequencing (ChIP-Seq) revealed that Zfp422 regulated ephrin type-A receptor 7 (EphA7) expression by binding an upstream 169-bp DNA sequence, which was proved to be an enhancer of EphA7. Knocking EphA7 down in C2C12 cells or deleting Zfp422 in myoblasts will inhibit cell apoptosis which is required for myoblast differentiation. These results indicate that Zfp422 is essential for skeletal muscle differentiation and fusion, through regulating EphA7 expression to maintain proper apoptosis.

Sign in / Sign up

Export Citation Format

Share Document