NDLI: Novel Split-Luciferase-Based Genetically Encoded Biosensors for Noninvasive Visualization of Rho GTPases

Content Provider	PubMed Central
Author	Leng, Weibing Pang, Xiaohui Xia, Hongwei Li, Mingxing Chen, Liu Tang, Qiulin Yuan, Dandan Ronghui, Li Libo, Li Gao, Fabao Feng, Bi
Editor	Aspenstrom, Pontus
Copyright Year	2013
Abstract	Rho family GTPases are critical regulators of many important cellular processes and the dysregulation of their activities is implicated in a variety of human diseases including oncogenesis and propagation of malignancy. The traditional methods, such as “pull-down” or two-hybrid procedures, are poorly suited to dynamically evaluate the activity of Rho GTPases, especially in living mammalian cells. To provide a novel alternative approach to analyzing Rho GTPase-associated signaling pathways in vivo, we developed a series of bioluminescent biosensors based on the genetically engineered firefly luciferase. These split-luciferase-based biosensors enable non-invasive visualization and quantification of the activity of Rho GTPases in living subjects. The strategy is to reasonably split the gene of firefly luciferase protein into two inactive fragments and then respectively fuse the two fragments to Rho GTPase and the GTPase-binding domain (GBD) of the specific effector. Upon Rho GTPase interacting with the binding domain in a GTP-dependent manner, these two luciferase fragments are brought into close proximity, leading to luciferase reconstitution and photon production in the presence of the substrate. Using these bimolecular luminescence complementation (BiLC) biosensors, we successfully visualized and quantified the activities of the three best characterized Rho GTPases by measuring the luminescence in living cells. We also experimentally investigated the sensitivity of these Rho GTPase biosensors to upstream regulatory proteins and extracellular ligands without lysing cells and doing labor-intensive works. By virtue of the unique functional characteristics of bioluminescence imaging, the BiLC-based biosensors provide an enormous potential for in vivo imaging of Rho GTPase signaling pathways and high-throughput screening of therapeutic drugs targeted to Rho GTPases and (or) upstream molecules in the near future.
Related Links	http://dx.doi.org/10.1371/journal.pone.0062230
Starting Page	62230
File Format	PDF
ISSN	19326203
e-ISSN	19326203
Journal	PLoS ONE
Issue Number	4
Volume Number	8
Language	English
Publisher	Public Library of Science
Publisher Date	2013-04-16
Access Restriction	Open
Rights Holder	Public Library of Science
Subject Keyword	Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

Sl.	Authority	Responsibilities	Communication Details
1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
5	Website/Portal (Helpdesk)	Queries regarding NDLI and its services	support@ndl.gov.in
6	Contents and Copyright Issues	Queries related to content curation and copyright issues	content@ndl.gov.in
7	National Digital Library of India Club (NDLI Club)	Queries related to NDLI Club formation, support, user awareness program, seminar/symposium, collaboration, social media, promotion, and outreach	clubsupport@ndl.gov.in
8	Digital Preservation Centre (DPC)	Assistance with digitizing and archiving copyright-free printed books	dpc@ndl.gov.in
9	IDR Setup or Support	Queries related to establishment and support of Institutional Digital Repository (IDR) and IDR workshops	idr@ndl.gov.in

Genetically encoded split luciferase biosensors to measure endosome disruption in real time in live cells

cAMP Biosensors Based on Genetically Encoded Fluorescent/Luminescent Proteins

Genetically Encoded Biosensors Based on Fluorescent Proteins

Multiplexed visualization of dynamic signaling networks using genetically encoded fluorescent protein-based biosensors

Genetically encoded biosensors based on engineered fluorescent proteins† ‡

A bacteria colony-based screen for optimal linker combinations in genetically encoded biosensors

Biosensors for Characterizing the Dynamics of Rho Family GTPases in Living Cells

Genetically Encoded Fluorescent Biosensors for Biomedical Applications

Visualization of ATP levels inside single living cells with fluorescence resonance energy transfer-based genetically encoded indicators

Novel Split-Luciferase-Based Genetically Encoded Biosensors for Noninvasive Visualization of Rho GTPases

Similar Documents

Genetically encoded split luciferase biosensors to measure endosome disruption in real time in live cells

cAMP Biosensors Based on Genetically Encoded Fluorescent/Luminescent Proteins

Genetically Encoded Biosensors Based on Fluorescent Proteins

Multiplexed visualization of dynamic signaling networks using genetically encoded fluorescent protein-based biosensors

Genetically encoded biosensors based on engineered fluorescent proteins† ‡

A bacteria colony-based screen for optimal linker combinations in genetically encoded biosensors

Biosensors for Characterizing the Dynamics of Rho Family GTPases in Living Cells

Genetically Encoded Fluorescent Biosensors for Biomedical Applications

Visualization of ATP levels inside single living cells with fluorescence resonance energy transfer-based genetically encoded indicators

Novel Split-Luciferase-Based Genetically Encoded Biosensors for Noninvasive Visualization of Rho GTPases