Mitochondrial uncouplers inhibit clathrin-mediated endocytosis largely through cytoplasmic acidification.
ATP production requires the establishment of an electrochemical proton gradient across the inner mitochondrial membrane. Mitochondrial uncouplers dissipate this proton gradient and disrupt numerous cellular processes, including vesicular trafficking, mainly through energy depletion. Here we show that Endosidin9 (ES9), a novel mitochondrial uncoupler, is a potent inhibitor of clathrin-mediated endocytosis (CME) in different systems and that ES9 induces inhibition of CME not because of its effect on cellular ATP, but rather due to its protonophore activity that leads to cytoplasm acidification. We show that the known tyrosine kinase inhibitor tyrphostinA23, which is routinely used to block CME, displays similar properties, thus questioning its use as a specific inhibitor of cargo recognition by the AP-2 adaptor complex via tyrosine motif-based endocytosis signals. Furthermore, we show that cytoplasm acidification dramatically affects the dynamics and recruitment of clathrin and associated adaptors, and leads to reduction of phosphatidylinositol 4,5-biphosphate from the plasma membrane.
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Wim
W
Dejonghe
VIB Center for the Biology of Disease, Laboratory of Neuronal Communication, 3000 Leuven, Belgium.|Department for Human Genetics, and Leuven Institute for Neurodegenerative Diseases, KU Leuven, 3000 Leuven, Belgium.
Sabine
S
Kuenen
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Evelien
E
Mylle
Institute of Science and Technology Austria, 3400 Klosterneuburg, Austria.
Mina
M
Vasileva
Department of Plant Physiology, Umeå Plant Science Centre, Umeå University, 90187 Umeå, Sweden.
Olivier
0000-0002-0546-7721
O
Keech
Department of Plant Physiology, Institute of Biochemistry and Biology, University of Potsdam, 14476 Potsdam, Germany.
Corrado
C
Viotti
VIB Center for the Biology of Disease, Laboratory of Neuronal Communication, 3000 Leuven, Belgium.|Department for Human Genetics, and Leuven Institute for Neurodegenerative Diseases, KU Leuven, 3000 Leuven, Belgium.
Jef
J
Swerts
Institute of Science and Technology Austria, 3400 Klosterneuburg, Austria.
Matyáš
M
Fendrych
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Fausto Andres
FA
Ortiz-Morea
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Kiril
K
Mishev
Developmental Biology of Plants, Centre for Organismal Studies, Heidelberg University, 69120 Heidelberg, Germany.
Simon
S
Delang
Developmental Biology of Plants, Centre for Organismal Studies, Heidelberg University, 69120 Heidelberg, Germany.
Stefan
S
Scholl
Department of Plant Cell Biology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1090 GE Amsterdam, The Netherlands.
Xavier
X
Zarza
Department of Cellular Biochemistry, Institute for Biochemistry and Biotechnology, Martin-Luther-University, 06120 Halle, Germany.
Mareike
M
Heilmann
Department of Plant Cell Biology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1090 GE Amsterdam, The Netherlands.
Jiorgos
J
Kourelis
VIB Center for the Biology of Disease, Laboratory of Neuronal Communication, 3000 Leuven, Belgium.|Department for Human Genetics, and Leuven Institute for Neurodegenerative Diseases, KU Leuven, 3000 Leuven, Belgium.
Jaroslaw
J
Kasprowicz
Compound Screening Facility, VIB, 9052 Ghent, Belgium.
Le Son Long
le SL
Nguyen
Compound Screening Facility, VIB, 9052 Ghent, Belgium.
Andrzej
A
Drozdzecki
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Isabelle
I
Van Houtte
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Anna-Mária
AM
Szatmári
Department of Forest Genetics and Plant Physiology, Umeå Plant Science Centre, Swedish University of Agricultural Sciences, 90183 Umeå, Sweden.
Mateusz
M
Majda
Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
Gary
G
Baisa
Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
Sebastian York
SY
Bednarek
Department of Forest Genetics and Plant Physiology, Umeå Plant Science Centre, Swedish University of Agricultural Sciences, 90183 Umeå, Sweden.
Stéphanie
S
Robert
Compound Screening Facility, VIB, 9052 Ghent, Belgium.
Dominique
D
Audenaert
Department of Plant Cell Biology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1090 GE Amsterdam, The Netherlands.
Christa
C
Testerink
Department of Plant Cell Biology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1090 GE Amsterdam, The Netherlands.
Teun
T
Munnik
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Daniël
D
Van Damme
Department of Cellular Biochemistry, Institute for Biochemistry and Biotechnology, Martin-Luther-University, 06120 Halle, Germany.
Ingo
I
Heilmann
Developmental Biology of Plants, Centre for Organismal Studies, Heidelberg University, 69120 Heidelberg, Germany.
Karin
K
Schumacher
Laboratory for Organic Synthesis, Department of Organic and Macromolecular Chemistry, Ghent University, 9000 Gent, Belgium.
Johan
J
Winne
Institute of Science and Technology Austria, 3400 Klosterneuburg, Austria.
Jiří
J
Friml
VIB Center for the Biology of Disease, Laboratory of Neuronal Communication, 3000 Leuven, Belgium.|Department for Human Genetics, and Leuven Institute for Neurodegenerative Diseases, KU Leuven, 3000 Leuven, Belgium.
Patrik
P
Verstreken
Department of Plant Systems Biology, VIB, 9052 Gent, Belgium.|Department of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Gent, Belgium.
Eugenia
E
Russinova
D000143:Acids / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000172:deficiency / Q000378:metabolism; D017360:Arabidopsis / Q000187:drug effects / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism*; D004705:Endocytosis / Q000187:drug effects*; D004734:Energy Metabolism / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D015388:Organelles / Q000187:drug effects / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D015363:Quinolones / Q000737:chemistry / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology*
D000143:Acids; D002966:Clathrin; D015363:Quinolones; D014475:Uncoupling Agents; D000255:Adenosine Triphosphate