Activation of the epithelial sodium channel by the metalloprotease meprin β subunit.
The Epithelial Na(+) Channel (ENaC) is an apical heteromeric channel that mediates Na(+) entry into epithelial cells from the luminal cell surface. ENaC is activated by proteases that interact with the channel during biosynthesis or at the extracellular surface. Meprins are cell surface and secreted metalloproteinases of the kidney and intestine. We discovered by affinity chromatography that meprins bind γ-ENaC, a subunit of the ENaC hetero-oligomer. The physical interaction involves NH(2)-terminal cytoplasmic residues 37-54 of γ-ENaC, containing a critical gating domain immediately before the first transmembrane domain, and the cytoplasmic COOH-terminal tail of meprin β (residues 679-704). This potential association was confirmed by co-expression and co-immunoprecipitation studies. Functional assays revealed that meprins stimulate ENaC expressed exogenously in Xenopus oocytes and endogenously in epithelial cells. Co-expression of ENaC subunits and meprin β or α/β in Xenopus oocytes increased amiloride-sensitive Na(+) currents approximately two-fold. This increase was blocked by preincubation with an inhibitor of meprin activity, actinonin. The meprin-mediated increase in ENaC currents in oocytes and epithelial cell monolayers required meprin β, but not the α subunit. Meprin β promoted cleavage of α and γ-ENaC subunits at sites close to the second transmembrane domain in the extracellular domain of each channel subunit. Thus, meprin β regulates the activity of ENaC in a metalloprotease-dependent fashion.
Cystic Fibrosis/Pulmonary Research and Treatment Center, Pennsylvania State University College of Medicine, Hershey, USA. agarciacaballero@yahoo.com
Agustin
A
Garcia-Caballero
Susan S
SS
Ishmael
Yan
Y
Dang
Daniel
D
Gillie
Judith S
JS
Bond
Sharon L
SL
Milgram
M Jackson
MJ
Stutts
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002846:Chromatography, Affinity; D004285:Dogs; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D047468:Immunoprecipitation; D015640:Ion Channel Gating; D007668:Kidney / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000494:pharmacology; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012964:Sodium / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D014981:Xenopus
D053503:Epithelial Sodium Channels; D006877:Hydroxamic Acids; D011480:Protease Inhibitors; D011994:Recombinant Proteins; C507048:SCNN1B protein, human; C507052:SCNN1G protein, human; C440894:Scnn1a protein, rat; C507050:Scnn1b protein, rat; C507054:Scnn1g protein, rat; D012964:Sodium; D008666:Metalloendopeptidases; C053833:meprin A; C010531:actinonin