Detail of Figure3C_20200717_4493NES_2nd_post_handBL



Project
Title
Timelapse images of the MDCK cysts without 2pabPAC activated by 1P excitation.
Description
Timelapse images of the MDCK cysts without 2pabPAC activated by 1P excitation.
Release, Updated
2022-03-31
License
CC-BY
Kind
Image data
File Formats
.oib
Data size
31.0 MB

Organism
Canis lupus familiaris ( NCBITaxon:9615 )
Strain(s)
-
Cell Line
MDCK cell ( CLO_0007646 )

Datatype
-
Molecular Function (MF)
-
Biological Process (BP)
cAMP-mediated signaling ( GO:0019933 )
Cellular Component (CC)
cAMP-dependent protein kinase complex ( GO:0005952 )
Biological Imaging Method
time lapse microscopy ( Fbbi:00000249 )
X scale
0.497 micrometer/pixel
Y scale
0.497 micrometer/pixel
Z scale
2 micrometer/pixel
T scale
20 sec per time interval

Image Acquisition
Experiment type
-
Microscope type
-
Acquisition mode
-
Contrast method
-
Microscope model
-
Detector model
-
Objective model
-
Filter set
-

Summary of Methods
See details in Kinjo T, et. al. (2020) ACS Chem Biol., 15(11):2848-2853.
Related paper(s)

Tomoaki Kinjo, Tetsuya Watabe, Kenju Kobachi, Kenta Terai, Michiyuki Matsuda (2020) Single-Cell Activation of the cAMP-Signaling Pathway in 3D Tissues with FRET-Assisted Two-Photon Activation of bPAC., ACS chemical biology, Volume 15, Number 11, pp. 2848-2853

Published in 2020 Nov 20 (Electronic publication in Oct. 19, 2020, midnight )

(Abstract) Bacterial photoactivated adenylyl cyclase (bPAC) has been widely used in signal transduction research. However, due to its low two-photon absorption, bPAC cannot be efficiently activated by two-photon (2P) excitation. Taking advantage of the high two-photon absorption of monomeric teal fluorescent protein 1 (mTFP1), we herein developed 2P-activatable bPAC (2pabPAC), a fusion protein consisting of bPAC and mTFP1. In 2pabPAC, the energy absorbed by mTFP1 excites bPAC by Furster resonance energy transfer (FRET) at ca. 43% efficiency. The light-induced increase in cAMP was monitored by a red-shifted FRET biosensor for PKA. In 3D MDCK cells and mouse liver, PKA was activated at single-cell resolution under a 2P microscope. We found that PKA activation in a single hepatocyte caused PKA activation in neighboring cells, indicating the propagation of PKA activation. Thus, 2pabPAC will provide a versatile platform for controlling the cAMP signaling pathway and investigating cell-to-cell communication in vivo.
(MeSH Terms)

Contact
Kenta Terai , Graduate School of Medicine, Kyoto University , Department of Pathology and Biology of Diseases
Contributors

OMERO Dataset
OMERO Project
Source