Cytochalasin E alters the cytoskeleton and decreases ENaC activity in Xenopus 2F3 cells.
Numerous reports have linked cytoskeleton-associated proteins with the regulation of epithelial Na(+) channel (ENaC) activity. The purpose of the present study was to determine the effect of actin cytoskeleton disruption by cytochalasin E on ENaC activity in Xenopus 2F3 cells. Here, we show that cytochalasin E treatment for 60 min can disrupt the integrity of the actin cytoskeleton in cultured Xenopus 2F3 cells. We show using single channel patch-clamp experiments and measurements of short-circuit current that ENaC activity, but not its density, is altered by cytochalasin E-induced disruption of the cytoskeleton. In nontreated cells, 8 of 33 patches (24%) had no measurable ENaC activity, whereas in cytochalasin E-treated cells, 17 of 32 patches (53%) had no activity. Analysis of those patches that did contain ENaC activity showed channel open probability significantly decreased from 0.081 ± 0.01 in nontreated cells to 0.043 ± 0.01 in cells treated with cytochalasin E. Transepithelial current from mpkCCD cells treated with cytochalasin E, cytochalasin D, or latrunculin B for 60 min was decreased compared with vehicle-treated cells. The subcellular expression of fodrin changed significantly, and several protein elements of the cytoskeleton decreased at least twofold after 60 min of cytochalasin E treatment. Cytochalasin E treatment disrupted the association between ENaC and myristoylated alanine-rich C-kinase substrate. The results presented here suggest disruption of the actin cytoskeleton by different compounds can attenuate ENaC activity through a mechanism involving changes in the subcellular expression of fodrin, several elements of the cytoskeleton, and destabilization of the ENaC-myristoylated alanine-rich C-kinase substrate complex.
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Matthew S
MS
Reifenberger
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Ling
L
Yu
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Hui-Fang
HF
Bao
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Billie Jeanne
BJ
Duke
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Bing-Chen
BC
Liu
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
He-Ping
HP
Ma
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia aalli@emory.edu.
Ahmed A
AA
Alli
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Douglas C
DC
Eaton
Department of Physiology and the Center for Cell and Molecular Signaling, Emory University School of Medicine, Atlanta, Georgia.
Abdel A
AA
Alli
D000199:Actins / Q000378:metabolism; D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D015638:Cytochalasin D / Q000378:metabolism; D003572:Cytochalasins / Q000494:pharmacology*; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D014981:Xenopus; D014982:Xenopus laevis; D013049:Spectrin
D000199:Actins; D020533:Angiogenesis Inhibitors; D002352:Carrier Proteins; D015638:Cytochalasin D; D003572:Cytochalasins; D053503:Epithelial Sodium Channels; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D013049:Spectrin; C007215:cytochalasin E
actin; cytoskeleton; epithelial Na+ channels