Data from: Calcium imaging with genetically encoded sensor Case12: facile analysis of α7/α9 nAChR mutants

Shelukhina I, Spirova E, Kudryavtsev D, Ojomoko L, Werner M, Methfessel C, Hollmann M, Tsetlin V

Date Published: August 11, 2017

DOI: http://dx.doi.org/10.5061/dryad.4k831

 

Files in this package

Content in the Dryad Digital Repository is offered "as is." By downloading files, you agree to the Dryad Terms of Service. To the extent possible under law, the authors have waived all copyright and related or neighboring rights to this data. CC0 (opens a new window) Open Data (opens a new window)

Title Photos 101-115
Downloaded 4 times
Description Photos 101-115 – Cytochemical fluorescent and bright field images were generated for quantification of Case12 and Alexa Fluor 555-α-bungarotoxin fluorescence in Neuro2a cells expressing human α7 nAChR (nicotinic acetylcholine receptor); Photos 201-210 – Cytochemical fluorescent and bright field images were generated for quantification of Alexa Fluor 555-α-bungarotoxin fluorescence and background fluorescence (in green channel) in Neuro2a cells expressing human α7 nAChR; Photos 301-316 – Cytochemical fluorescent and bright field images were generated for quantification of Case12 fluorescence and background fluorescence (in red channel) in Neuro2a cells expressing human α7 nAChR; Photos 401-413 – Cytochemical fluorescent and bright field images were generated for quantification of background fluorescence (in green and red channels) in Neuro2a cells expressing human α7 nAChR; Photos 501-505 – Cytochemical fluorescent and bright field images were generated for quantification of Case12 and Alexa Fluor 555-α-bungarotoxin fluorescence in Neuro2a cells expressing mouse muscle (WT nAChR; Photos 604-612 – Cytochemical fluorescent and bright field images were generated for quantification of Alexa Fluor 555-α-bungarotoxin fluorescence and background fluorescence (in green channel) in Neuro2a cells expressing mouse muscle (WT) nAChR; Photos 701-710 – Cytochemical fluorescent and bright field images were generated for quantification of TMRE (Tetramethylrhodamine, ethyl ester; 20 nM) labeling of Neuro2a cells expressing human α7 nAChR; Photos 801-805 – Cytochemical fluorescent and bright field images were generated for quantification of PI (Propidium iodide; 50 ng/ml) labeling of Neuro2a cells expressing human α7 nAChR
Download README.docx (29.33 Kb)
Download Cytochemical photos 101-805_Alexa555-aBgt...PI.zip (109.8 Mb)
Details View File Details
Title Video_Calcium response of N2A cells_a7 nAChR
Downloaded 4 times
Description Videos show calcium response of Neuro2a cells expressing human α7 nAChR (nicotinic acetylcholine receptor), a chaperone NACHO and a fluorescent genetically-encoded calcium sensor Case12 to different concentration of acetylcholine in the presence of a positive allosteric modulator PNU120596. Controls in the presence of α-cobratoxin are presented as well.
Download Video_Calcium response of N2A cells_a7 nAChR.zip (170.4 Mb)
Download README.docx (28.2 Kb)
Details View File Details
Title Electrophysiology and Ca_imaging normalized data
Downloaded 2 times
Description The file contains the normalized electrophysiological (TEVC (two-electrode voltage clamp)) and calcium imaging (Ca-img) data for oocyte and Neuro2a cells responses, respectively. WT human α7 nAChR (nicotinic acetylcholine receptor) and mouse muscle nAChR were expressed in oocytes and Neuro2a cells. Besides, their mutants (at positions 117-119, 184, 185, 187 and 189 in α7 nAChR and at positions 153 and 190 in muscle nAChR) were heterologously expressed as well and their responses were estimated accordingly. Responses to different concentrations of acetylcholine (Ach) and epibatidine (Epi) were measured. For calcium imaging studies two calcium sensors (genetically encoded Case12 and low-molecular weight Fluo-4) were used. Calcium imaging analysis was carried out in the presence of a positive allosteric modulator PNU120596.
Download Electrophysiology and Ca_imaging normalize....xlsx (64.98 Kb)
Details View File Details
Title pdb
Downloaded 2 times
Description PDB file WT.pdb contains the last frame from 10 ns molecular dynamics simulation alpha7-AChBP chimera (PDB 3SQ6). Only two adjacent subunits were subjected to simulations. PDB files with names E185V, E189G, F187S, L119D, Q117T, R186I, S184N and Y118W contain the last frame from molecular dynamics simulation of alpha7-AChBP chimera with the respective mutations (residues numbered according to human alpha7 nAChR). PDB files F104.pdb and V104_mutant.pdb contain structures in which 104th residue of alpha7 AChBP chimera was changed either to phenylalanine (as in actual human alpha7 nAChR) or to valine (as in alpha9 nAChR).
Download pdb.zip (1.063 Mb)
Details View File Details

When using this data, please cite the original publication:

Shelukhina I, Spirova E, Kudryavtsev D, Ojomoko L, Werner M, Methfessel C, Hollmann M, Tsetlin V (2017) Calcium imaging with genetically encoded sensor Case12: Facile analysis of α7/α9 nAChR mutants. PLOS ONE 12(8): e0181936. http://dx.doi.org/10.1371/journal.pone.0181936

Additionally, please cite the Dryad data package:

Shelukhina I, Spirova E, Kudryavtsev D, Ojomoko L, Werner M, Methfessel C, Hollmann M, Tsetlin V (2017) Data from: Calcium imaging with genetically encoded sensor Case12: facile analysis of α7/α9 nAChR mutants. Dryad Digital Repository. http://dx.doi.org/10.5061/dryad.4k831
Cite | Share
Download the data package citation in the following formats:
   RIS (compatible with EndNote, Reference Manager, ProCite, RefWorks)
   BibTex (compatible with BibDesk, LaTeX)

Search for data

Be part of Dryad

We encourage organizations to: