Distinct roles for luminal acidification in apical protein sorting and trafficking in zebrafish

Epithelial cell physiology critically depends on the asymmetric distribution of channels and transporters. However, the mechanisms targeting membrane proteins to the apical surface are still poorly understood. Here, we performed a visual forward genetic screen in the zebrafish intestine and identifi...

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Published inThe Journal of cell biology Vol. 219; no. 4; p. 1
Main Authors Levic, Daniel S, Ryan, Sean, Marjoram, Lindsay, Honeycutt, Jamie, Bagwell, Jennifer, Bagnat, Michel
Format Journal Article
LanguageEnglish
Published United States Rockefeller University Press 06.04.2020
Subjects
Acidification
Adenosine triphosphatase
Animals
Danio rerio
Development
Epithelial cells
Genetic screening
Golgi apparatus
H+-transporting ATPase
Hydrogen
Hydrogen-Ion Concentration
Intestine
Low temperature
Membrane proteins
Membrane Proteins - genetics
Membrane Proteins - metabolism
Membranes
Mutants
Mutation
Perturbation
Phenobarbital - chemistry
Phenobarbital - metabolism
Physiology
Polarity
Protein Transport
Proteins
Proton-Translocating ATPases - genetics
Proton-Translocating ATPases - metabolism
Skewed distributions
Sodium
Trafficking
Transport
Zebrafish
Zebrafish - metabolism
Zebrafish Proteins - genetics
Zebrafish Proteins - metabolism
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Summary:Epithelial cell physiology critically depends on the asymmetric distribution of channels and transporters. However, the mechanisms targeting membrane proteins to the apical surface are still poorly understood. Here, we performed a visual forward genetic screen in the zebrafish intestine and identified mutants with defective apical targeting of membrane proteins. One of these mutants, affecting the vacuolar H+-ATPase gene atp6ap1b, revealed specific requirements for luminal acidification in apical, but not basolateral, membrane protein sorting and transport. Using a low temperature block assay combined with genetic and pharmacologic perturbation of luminal pH, we monitored transport of newly synthesized membrane proteins from the TGN to apical membrane in live zebrafish. We show that vacuolar H+-ATPase activity regulates sorting of O-glycosylated proteins at the TGN, as well as Rab8-dependent post-Golgi trafficking of different classes of apical membrane proteins. Thus, luminal acidification plays distinct and specific roles in apical membrane biogenesis.
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S. Ryan’s present address is Department of Biology, St. Bonaventure University, St. Bonaventure, NY.
ISSN:0021-9525
1540-8140
DOI:10.1083/jcb.201908225