scholarly journals Cell Type-specific Protein-DNA Interactions at the cAMP Response Elements of the Prohormone Convertase 1 Promoter

1997 ◽  
Vol 272 (4) ◽  
pp. 2500-2508 ◽  
Author(s):  
Erik Jansen ◽  
Torik A. Y. Ayoubi ◽  
Sandra M. P. Meulemans ◽  
Wim J. M. Van de Ven
Keyword(s):  
Specific Protein ◽  
Prohormone Convertase ◽  
Cell Type ◽  
Dna Interactions ◽  
Response Elements ◽  
Prohormone Convertase 1 ◽  
Protein Dna Interactions ◽  
Cell Type Specific

scholarly journals FlyORF-TaDa allows rapid generation of new lines for in vivo cell-type specific profiling of protein-DNA interactions in Drosophila melanogaster

2020 ◽  
Author(s):  
Gabriel N Aughey ◽  
Caroline Delandre ◽  
Tony D Southall ◽  
Owen J Marshall
Keyword(s):  
Open Reading Frames ◽  
Cell Type ◽  
Dna Interactions ◽  
Active Chromatin ◽  
Protein Dna Interactions ◽  
Genetic Cross ◽  
Cell Type Specific ◽  
Rapid Generation ◽  
Reading Frames

AbstractTargeted DamID (TaDa) is an increasingly popular method of generating cell-type specific DNA binding profiles in vivo. Although sensitive and versatile, TaDa requires the generation of new transgenic fly lines for every protein that is profiled, which is both time-consuming and costly. Here, we describe the FlyORF-TaDa system for converting an existing FlyORF library of inducible open reading frames (ORFs) to TaDa lines via a genetic cross, with recombinant progeny easily identifiable by eye colour. Profiling the binding of the H3K36me3-associated chromatin protein MRG15 in larval neural stem cells using both FlyORF-TaDa and conventional TaDa demonstrates that new lines generated using this system provide accurate and highly-reproducible DamID binding profiles. Our data further show that MRG15 binds to a subset of active chromatin domains in vivo. Courtesy of the large coverage of the FlyORF library, the FlyORF-TaDa system enables the easy creation of TaDa lines for 74% of all transcription factors and chromatin modifying proteins within the Drosophila genome.

scholarly journals Membrane-bound GFP-labelled vectors for Targeted DamID allow simultaneous profiling of expression domains and DNA binding

2020 ◽  
Author(s):  
Caroline Delandre ◽  
John PD McMullen ◽  
Owen J Marshall
Keyword(s):  
Translational Control ◽  
Expression Patterns ◽  
Insertion Site ◽  
Promoter Strength ◽  
Cell Type ◽  
Dna Interactions ◽  
Cell Type Specificity ◽  
Antibody Staining ◽  
Protein Dna Interactions ◽  
Cell Type Specific

AbstractTargeted DamID (TaDa) allows highly efficient cell-type-specific profiling of protein-DNA interactions. Cell-type-specificity, however, is governed by the GAL4/UAS system, which can exhibit differences in expression patterns depending upon the genomic insertion site and the UAS promoter strength. The TaDa system uses a bicistronic transcript to reduce the translation rates of Dam-fusion proteins, presenting the possibility of using the primary ORF within in the transcript to label expression domains and precisely identified the profiled cell populations in experimental samples. Here, we describe two TaDa vectors, pTaDaG and pTaDaG2, that use myristoylated GFP as the primary ORF. Differing lengths of the myristoylation sequence between the plasmids allows additional translational control. Fly lines created with this system allow easy visualisation of expression domains under both fluorescent dissecting and confocal microscopes without the use of antibody staining, whilst faithfully profiling protein-DNA interactions via Targeted DamID.

scholarly journals Generating Cell Type-Specific Protein Signatures from Non-symptomatic and Diseased Tissues

2020 ◽  
Vol 48 (8) ◽  
pp. 2218-2232
Author(s):  
Jessica S. Sadick ◽  
Lorin A. Crawford ◽  
Harry C. Cramer ◽  
Christian Franck ◽  
Shane A. Liddelow ◽  
...  
Keyword(s):  
Specific Protein ◽  
Cell Type ◽  
Protein Signatures ◽  
Cell Type Specific

scholarly journals Cell Type-Specific Replication of Simian Virus 40 Conferred by Hormone Response Elements in the Late Promoter

2002 ◽  
Vol 76 (13) ◽  
pp. 6762-6770 ◽  
Author(s):  
Michael L. Farrell ◽  
Janet E. Mertz
Keyword(s):  
Simian Virus 40 ◽  
Simian Virus ◽  
Genome Replication ◽  
Hormone Response ◽  
Cell Type ◽  
Late Promoter ◽  
African Green Monkey Kidney ◽  
Response Elements ◽  
Hormone Response Elements ◽  
Cell Type Specific

ABSTRACT The late genes of SV40 are not expressed at significant levels until after the onset of viral DNA replication. We previously identified two hormone response elements (HREs) in the late promoter that contribute to this delay. Mutants defective in these HREs overexpress late RNA at early, but not late, times after transfection of CV-1PD cells. Overexpression of nuclear receptors (NRs) that recognize these HREs leads to repression of the late promoter in a sequence-specific and titratable manner, resulting in a delay in late gene expression. These observations led to a model in which the late promoter is repressed at early times after infection by NRs, with this repression being relieved by titration of these repressors through simian virus 40 (SV40) genome replication to high copy number. Here, we tested this model in the context of the viral life cycle. SV40 genomes containing mutations in either or both HREs that significantly reduce NR binding without altering the coding of any proteins were constructed. Competition for replication between mutant and wild-type viruses in low-multiplicity coinfections indicated that the +1 HRE offered a significant selective advantage to the virus within a few cycles of infection in African green monkey kidney cell lines CV-1, CV-1P, TC-7, MA-134, and Vero but not in CV-1PD′ cells. Interestingly, the +55 HRE offered a selective disadvantage in MA-134 cells but had no effect in CV-1, CV-1P, TC-7, Vero, and CV-1PD′ cells. Thus, we conclude that these HREs are biologically important to the virus, but in a cell type-specific manner.

scholarly journals Btbd11 is an inhibitory interneuron specific synaptic scaffolding protein that supports excitatory synapse structure and function

2021 ◽  
Author(s):  
Alexei M. Bygrave ◽  
Ayesha Sengupta ◽  
Ella P. Jackert ◽  
Mehroz Ahmed ◽  
Beloved Adenuga ◽  
...  
Keyword(s):  
Inhibitory Interneuron ◽  
Nmda Receptor Antagonist ◽  
Specific Protein ◽  
Network Activity ◽  
Scaffolding Protein ◽  
Inhibitory Interneurons ◽  
Cell Type ◽  
Cell Type Specific

Synapses in the brain exhibit cell–type–specific differences in basal synaptic transmission and plasticity. Here, we evaluated cell–type–specific differences in the composition of glutamatergic synapses, identifying Btbd11, as an inhibitory interneuron–specific synapse–enriched protein. Btbd11 is highly conserved across species and binds to core postsynaptic proteins including Psd–95. Intriguingly, we show that Btbd11 can undergo liquid–liquid phase separation when expressed with Psd–95, supporting the idea that the glutamatergic post synaptic density in synapses in inhibitory and excitatory neurons exist in a phase separated state. Knockout of Btbd11 from inhibitory interneurons decreased glutamatergic signaling onto parvalbumin–positive interneurons. Further, both in vitro and in vivo, we find that Btbd11 knockout disrupts network activity. At the behavioral level, Btbd11 knockout from interneurons sensitizes mice to pharmacologically induced hyperactivity following NMDA receptor antagonist challenge. Our findings identify a cell–type–specific protein that supports glutamatergic synapse function in inhibitory interneurons–with implication for circuit function and animal behavior.

scholarly journals In vivo protein-DNA interactions at the c-jun promoter: preformed complexes mediate the UV response.

1993 ◽  
Vol 13 (9) ◽  
pp. 5490-5499 ◽  
Author(s):  
D Rozek ◽  
G P Pfeifer
Keyword(s):  
Cpg Island ◽  
Uv Light ◽  
Dimethyl Sulfate ◽  
Related Factor ◽  
Dna Interactions ◽  
Intact Cells ◽  
Response Elements ◽  
Protein Dna Interactions ◽  
Uv Response

Irradiation of cells with UV light triggers a genetic response, called the UV response, which results in induction of a set of genes containing AP-1-binding sites. The c-jun gene itself, which codes for AP-1-binding activity, is strongly (> 100-fold) and rapidly activated by UV. The UV induction of c-jun is mediated by two UV response elements consisting of AP-1-like sequences within its 5' control region. We have analyzed protein-DNA interactions in vivo at the c-jun promoter in noninduced and UV-irradiated HeLa cells. In vivo footprint analysis was performed by using dimethyl sulfate on intact cells and DNase I on lysolecithihin-permeabilized cells in conjunction with ligation-mediated polymerase chain reaction to cover about 450 bp of the c-jun promoter, including the transcription start sites. We find that this region does not contain methylated cytosines and is thus a typical CpG island. In uninduced cells, in vivo protein-DNA interactions were localized to an AP-1-like sequence (nucleotides [nt] -71 to -64), a CCAAT box element (nt -91 to -87), two SP1 sequences (nt -115 to -110 and -123 to -118), a nuclear factor jun site (nt -140 to -132), and a second AP-1-like sequence (nt -190 to -183). These results indicate that complex protein-DNA interactions exist at the c-jun promoter prior to induction by an external stimulus. Surprisingly, after stimulation of c-jun expression by UV irradiation, all in vivo protein-DNA contacts remained essentially unchanged, including the two UV response elements located at the AP-1-like sequences. The UV-induced signalling cascade leads to phosphorylation of c-Jun on serines 63 and 73 (Y. Devary, R.A. Gottlieb, T. Smeal, and M. Karin, Cell 71:1081-1091, 1992). Taken together, these data suggest that modification of the transactivating domain of DNA-bound c-Jun or a closely related factor may trigger the rapid induction of the c-jun gene.

Sign in / Sign up

Export Citation Format

Share Document