Generation of Rab-based transgenic lines for in vivo studies of endosome biology in zebrafish. Dev Dyn 2011 Nov;240(11):2452-65
Date
10/07/2011Pubmed ID
21976318Pubmed Central ID
PMC3197968DOI
10.1002/dvdy.22758Scopus ID
2-s2.0-80054832731 (requires institutional sign-in at Scopus site) 75 CitationsAbstract
The Rab family of small GTPases function as molecular switches regulating membrane and protein trafficking. Individual Rab isoforms define and are required for specific endosomal compartments. To facilitate in vivo investigation of specific Rab proteins, and endosome biology in general, we have generated transgenic zebrafish lines to mark and manipulate Rab proteins. We also developed software to track and quantify endosome dynamics within time-lapse movies. The established transgenic lines ubiquitously express EGFP fusions of Rab5c (early endosomes), Rab11a (recycling endosomes), and Rab7 (late endosomes) to study localization and dynamics during development. Additionally, we generated UAS-based transgenic lines expressing constitutive active (CA) and dominant-negative (DN) versions for each of these Rab proteins. Predicted localization and functional consequences for each line were verified through a variety of assays, including lipophilic dye uptake and Crumbs2a localization. In summary, we have established a toolset for in vivo analyses of endosome dynamics and functions.
Author List
Clark BS, Winter M, Cohen AR, Link BAAuthor
Brian A. Link PhD Professor in the Cell Biology, Neurobiology and Anatomy department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
Amino Acid SequenceAnimals
Animals, Genetically Modified
Biology
Endosomes
Gene Transfer Techniques
Green Fluorescent Proteins
Membrane Proteins
Models, Biological
Neuroepithelial Cells
Sequence Homology, Amino Acid
Zebrafish
Zebrafish Proteins
rab GTP-Binding Proteins
rab5 GTP-Binding Proteins
