Transcriptional regulation of the Hedgehog effector CI by the zinc-finger gene combgap

Development ◽  
2000 ◽  
Vol 127 (19) ◽  
pp. 4095-4103 ◽  
Author(s):  
G.L. Campbell ◽  
A. Tomlinson
Keyword(s):  
Transcription Factors ◽  
Cell Fate ◽  
Zinc Finger ◽  
Transcriptional Activation ◽  
Dna Binding Protein ◽  
Loss Of Function ◽  
Cubitus Interruptus ◽  
Anterior Compartment ◽  
Zinc Finger Gene ◽  
Animal Development

Members of the Hedgehog (HH) family of secreted signaling molecules specify cell fate during animal development by controlling the activity of members of the Gli family of zinc-finger transcription factors in responding cells. In Drosophila the Gli homolog, cubitus interruptus (CI), is expressed only in the anterior compartment where it represses targets such as the signaling molecule genes decapentaplegic (dpp) and wingless (wg). HH is expressed in the posterior and diffuses into the anterior where it antagonizes CI repression resulting in dpp and wg expression immediately anterior to the compartment border. Reducing CI levels results in misexpression of wg and dpp, while CI misexpression in the posterior disrupts differentiation. Thus, normal disc patterning requires high levels of CI in the anterior and the absence of CI in the posterior. Here we show that mutations in combgap (cg) result in deregulation of CI expression, which is now expressed at much lower levels and ubiquitously, i.e., also in the posterior. Consequently, cg mutants phenocopy ci loss-of-function mutants in the anterior and ci gain-of-function mutants in the posterior. cg encodes a putative DNA-binding protein that regulates both transcriptional activation and repression of the ci gene.

Single zinc-finger extension: enhancing transcriptional activity and specificity of three-zinc-finger proteins

2005 ◽  
Vol 386 (2) ◽  
pp. 95-99 ◽  
Author(s):  
Alexander E.F. Smith ◽  
Farzin Farzaneh ◽  
Kevin G. Ford
Keyword(s):  
Transcription Factors ◽  
Fusion Protein ◽  
Zinc Finger ◽  
Protein Interactions ◽  
Transcriptional Activation ◽  
Zinc Finger Protein ◽  
Zinc Finger Proteins ◽  
Finger Protein ◽  
Finger Extension ◽  
Vp16 Fusion

AbstractIn order to demonstrate that an existing zinc-finger protein can be simply modified to enhance DNA binding and sequence discrimination in both episomal and chromatin contexts using existing zinc-finger DNA recognition code data, and without recourse to phage display and selection strategies, we have examined the consequences of a single zinc-finger extension to a synthetic three-zinc-finger VP16 fusion protein, on transcriptional activation from model target promoters harbouring the zinc-finger binding sequences. We report a nearly 10-fold enhanced transcriptional activation by the four-zinc-finger VP16 fusion protein relative to the progenitor three-finger VP16 protein in transient assays and a greater than five-fold enhancement in stable reporter-gene expression assays. A marked decrease in transcriptional activation was evident for the four-zinc-finger derivative from mutated regulatory regions compared to the progenitor protein, as a result of recognition site-size extension. This discriminatory effect was shown to be protein concentration-dependent. These observations suggest that four-zinc-finger proteins are stable functional motifs that can be a significant improvement over the progenitor three-zinc-finger protein, both in terms of specificity and the ability to target transcriptional function to promoters, and that single zinc-finger extension can therefore have a significant impact on DNA zinc-finger protein interactions. This is a simple route for modifying or enhancing the binding properties of existing synthetic zinc-finger-based transcription factors and may be particularly suited for the modification of endogenous zinc-finger transcription factors for promoter biasing applications.

Studying the consequences of immediate loss of gene function in the intestine: APC

2005 ◽  
Vol 33 (4) ◽  
pp. 665-666 ◽  
Author(s):  
A.R. Clarke
Keyword(s):  
Cell Fate ◽  
Gene Function ◽  
Transcriptional Activation ◽  
Underlying Disease ◽  
Loss Of Function ◽  
Critical Determinant ◽  
Proliferation And Apoptosis ◽  
Intestinal Tumour ◽  
Early Effects ◽  
Tumour Initiation

The use of mouse models to study neoplasia is proving particularly powerful in dissecting the mechanisms underlying disease initiation and progression. However, the majority of these models have been somewhat limited in studying the very early effects of loss of gene function, as tumour initiation relies upon either constitutive loss of gene function or spontaneous somatic loss of function. We have therefore adopted a strategy of using an inducible Cre-lox-based system to analyse the effects of loss of gene function, the use of which is reviewed here for the intestinal tumour suppressor APC (adenomatous polyposis coli). Using this approach, we have conditionally and synchronously inactivated APC in virtually all the epithelial cells of the adult murine small intestine. After 5 days following induction of Cre-mediated recombination, mice show grossly altered crypt/villus architecture. Deficiency in APC perturbs migration, alters the normal programme of differentiation and results in increased proliferation and apoptosis. Microarray analysis reveals the transcriptome to be significantly altered; reflecting both gross phenotypic changes and changes in transcriptional activation. These findings demonstrate that APC is indeed the critical determinant of cell fate in the intestinal epithelium, explaining its role as the cellular ‘gatekeeper’ in preventing neoplasia.

scholarly journals A cohort of Caenorhabditis species lacking the highly conserved let-7 microRNA

2020 ◽  
Author(s):  
Charles Nelson ◽  
Victor Ambros
Keyword(s):  
Cell Fate ◽  
Nematode Species ◽  
Loss Of Function ◽  
Animal Development ◽  
C Elegans ◽  
Adult Transition ◽  
Developmental Progression ◽  
Caenorhabditis Species ◽  
Heterochronic Gene ◽  
Bilaterian Animal

ABSTRACTlet-7 is a highly conserved microRNA with critical functions integral to cell fate specification and developmental progression in diverse animals. In Caenorhabditis elegans, let-7 is a component of the heterochronic (developmental timing) gene regulatory network, and loss-of-function mutations of let-7 result in lethality during the larval to adult transition due to misregulation of the conserved let-7 target, lin-41. To date, no bilaterian animal lacking let-7 has been characterized. In this study, we identify a cohort of nematode species within the genus Caenorhabditis, closely related to C. elegans, that lack the let-7 microRNA, owing to absence of the let-7 gene. Using C. sulstoni as a representative let-7-lacking species to characterize normal larval development in the absence of let-7, we demonstrate that, except for the lack of let-7, the heterochronic gene network is otherwise functionally conserved. We also report that species lacking let-7 contain a group of divergent let-7 orthologs -- also known as the let-7-family of microRNAs -- that have apparently assumed the role of targeting the lin-41 mRNA.Summary StatementWe have identified a group of Caenorhabditis species that lack let-7a, an otherwise highly conserved and nearly ubiquitous microRNA that was previously thought to be critical to bilaterian animal development.

scholarly journals Characterization of an E1A-CBP Interaction Defines a Novel Transcriptional Adapter Motif (TRAM) in CBP/p300

1999 ◽  
Vol 73 (5) ◽  
pp. 3574-3581 ◽  
Author(s):  
Mark J. O’Connor ◽  
Holger Zimmermann ◽  
Søren Nielsen ◽  
Hans-Ulrich Bernard ◽  
Tony Kouzarides
Keyword(s):  
Transcription Factors ◽  
Cell Fate ◽  
Transcriptional Activation ◽  
Molecular Model ◽  
P53 Protein ◽  
Small Region ◽  
Adenovirus E1a ◽  
Cellular Transcription

ABSTRACT The adenovirus E1A protein subverts cellular processes to induce mitotic activity in quiescent cells. Important targets of E1A include members of the transcriptional adapter family containing CBP/p300. Competition for CBP/p300 binding by various cellular transcription factors has been suggested as a means of integrating different signalling pathways and may also represent a potential mechanism by which E1A manipulates cell fate. Here we describe the characterization of the interaction between E1A and the C/H3 region of CBP. We define a novel conserved 12-residue transcriptional adapter motif (TRAM) within CBP/p300 that represents the binding site for both E1A and numerous cellular transcription factors. We also identify a sequence (FPESLIL) within adenovirus E1A that is required to bind the CBP TRAM. Furthermore, an E1A peptide containing the FPESLIL sequence is capable of preventing the interaction between CBP and TRAM-binding transcription factors, such as p53, E2F, and TFIIB, thus providing a molecular model for E1A action. As an in vivo demonstration of this model, we used a small region of CBP containing a functional TRAM that can bind to the p53 protein. The CBP TRAM binds p53 sequences targeted by the cellular regulator MDM2, and we demonstrate that an MDM2-p53 interaction can be disrupted by the CBP TRAM, leading to stabilization of cellular p53 levels and the activation of p53-dependent transcription. Transcriptional activation of p53 by the CBP TRAM is abolished by wild-type E1A but not by a CBP-binding-deficient E1A mutant.

scholarly journals E2F/Dp inactivation in fat body cells triggers systemic metabolic changes

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Maria Paula Zappia ◽  
Ana Guarner ◽  
Nadia Kellie-Smith ◽  
Alice Rogers ◽  
Robert Morris ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Cell Fate ◽  
Fat Body ◽  
Critical Role ◽  
Systemic Effects ◽  
Proteomic Profiling ◽  
Animal Development ◽  
Metabolomic Profiling ◽  
Essential Function ◽  
E2f Transcription Factors

The E2F transcription factors play a critical role in controlling cell fate. In Drosophila, the inactivation of E2F in either muscle or fat body results in lethality, suggesting an essential function for E2F in these tissues. However, the cellular and organismal consequences of inactivating E2F in these tissues are not fully understood. Here, we show that the E2F loss exerts both tissue-intrinsic and systemic effects. The proteomic profiling of E2F-deficient muscle and fat body revealed that E2F regulates carbohydrate metabolism, a conclusion further supported by metabolomic profiling. Intriguingly, animals with E2F-deficient fat body had a lower level of circulating trehalose and reduced storage of fat. Strikingly, a sugar supplement was sufficient to restore both trehalose and fat levels, and subsequently, rescued animal lethality. Collectively, our data highlight the unexpected complexity of E2F mutant phenotype, which is a result of combining both tissue-specific and systemic changes that contribute to animal development.

scholarly journals The ZNF35 human zinc finger gene encodes a sequence-specific DNA-binding protein

FEBS Letters ◽  
1993 ◽  
Vol 321 (2-3) ◽  
pp. 233-236 ◽  
Author(s):  
Gina Pengue ◽  
Paola Cannada-Bartoli ◽  
Luigi Lania
Keyword(s):  
Dna Binding ◽  
Zinc Finger ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
Zinc Finger Gene

engrailed and polyhomeotic interactions are required to maintain the A/P boundary of the Drosophila developing wing

Development ◽  
1998 ◽  
Vol 125 (15) ◽  
pp. 2771-2780 ◽  
Author(s):  
F. Maschat ◽  
N. Serrano ◽  
N.B. Randsholt ◽  
G. Geraud
Keyword(s):  
Cell Fate ◽  
Regulatory Protein ◽  
Polycomb Group ◽  
Posterior Compartment ◽  
Cubitus Interruptus ◽  
Anterior Compartment ◽  
Embryonic Segmentation ◽  
Wing Morphogenesis ◽  
Anterior Posterior

Engrailed is a nuclear regulatory protein with essential roles in embryonic segmentation and wing morphogenesis. One of its regulatory targets in embryos was shown to be the Polycomb group gene, polyhomeotic. We show here that transheterozygous adult flies, mutant for both engrailed and polyhomeotic, show a gap in the fourth vein. In the corresponding larval imaginal discs, a polyhomeotic-lacZ enhancer trap is not normally activated in anterior cells adjacent to the anterior-posterior boundary. This intermediary region corresponds to the domain of low engrailed expression that appears in the anterior compartment, during L3. Several arguments show that engrailed is responsible for the induction of polyhomeotic in these cells. The role of polyhomeotic in this intermediary region is apparently to maintain the repression of hedgehog in the anterior cells abutting the anterior-posterior boundary, since these cells ectopically express hedgehog when polyhomeotic is not activated. This leads to ectopic expressions first of patched, then of cubitus interruptus and decapentaplegic in the posterior compartment, except for the dorsoventral border cells that are not affected. Thus posterior cells express a new set of genes that are normally characteristic of anterior cells, suggesting a change in the cell identity. Altogether, our data indicate that engrailed and polyhomeotic interactions are required to maintain the anterior-posterior boundary and the posterior cell fate, just prior to the evagination of the wing.

scholarly journals Identification and expression of the cDNA of KIN17, a zinc-finger gene located on mouse chromosome 2, encoding a new DNA-binding protein

1991 ◽  
Vol 19 (19) ◽  
pp. 5117-5123 ◽  
Author(s):  
Jaime F. Angulo ◽  
Evelyne Rouer ◽  
Alexander Mazin ◽  
Marie-geneviève Mattei ◽  
Agnèves Tissier ◽  
...  
Keyword(s):  
Dna Binding ◽  
Mouse Chromosome ◽  
Zinc Finger ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
Chromosome 2 ◽  
Zinc Finger Gene ◽  
Mouse Chromosome 2

scholarly journals EPEN-05. MUTATIONAL ANALYSIS OF THE C11ORF95 DOMAIN AND SINGLE-CELL RNA-SEQ PROFILE OF A MOUSE MODEL OF SUPRATENTORIAL EPENDYMOMA

2021 ◽  
Vol 23 (Supplement_1) ◽  
pp. i14-i14
Author(s):  
Kevin Truong ◽  
James He ◽  
Gavin Birdsall ◽  
Ericka Randazzo ◽  
Jesse Dunnack ◽  
...  
Keyword(s):  
Mouse Model ◽  
Single Cell ◽  
Tumor Cells ◽  
Cell Fate ◽  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Cell Types ◽  
Marker Genes ◽  
Rna Seq ◽  
Loss Of Function

Abstract We used a recently developed mouse model to better understand the cellular and molecular determinants of tumors driven by the oncogenic fusion protein C11orf95-RELA. Our approach makes use of in utero electroporation and a binary transposase system to introduce human C11orf95-RELA sequence, wild type and mutant forms, into neural progenitors. We used single cell RNA-seq to profile the cellular constituents within the resulting tumors in mice. We find that approximately 70% of the cells in the tumors do not express the oncogene C11orf95-RELA and these non-oncogene expressing cells are a combination of different non-tumor cell cell-types, including significant numbers of T-cells, and macrophages. The C11orf95-RELA expressing tumor cells have a unique transcriptomic profile that includes both astrocytic and neural progenitor marker genes, and is distinct from glioblastoma transcriptomic profiles. Since C11orf95-RELA is believed to function through a combination of both activation of NF-κB response genes by constitutive activation of RELA, and genes not activated by NF-κB, we assessed the expression of NF-κB response genes across the populations of cells in the tumor. Interestingly, when tumor cells highly expressing C11orf95-RELA were analyzed further, the subclusters identified were distinguished by upregulation of non-NF-kB pathways involved in cell proliferation, cell fate determination, and immune activation. We hypothesized that the C11orf95 domain may function to bring RELA transcriptional activation to inappropriate non-NF-κB targets, and we therefore performed a point mutation analysis of the C11orf95 domain. We found that mutations in either of the cysteines or histidines that make up a possible zinc finger domain in C11orf95 eliminate the ability of the fusion to induce tumors. In cell lines, these loss-of-function point mutants still trafficked to nuclei, and activated NF-κB pathways. We are currently using RNAseq and CRISPR loss-of function to identify genes downstream of C11orf95-RELA that are required for tumorigenesis.

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