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Prostasin interacts with the epithelial Na+ channel and facilitates cleavage of
the ?-subunit by a second protease
#MMPMID25209858
Carattino MD
; Mueller GM
; Palmer LG
; Frindt G
; Rued AC
; Hughey RP
; Kleyman TR
Am J Physiol Renal Physiol
2014[Nov]; 307
(9
): F1080-7
PMID25209858
show ga
During maturation, the ?- and ?-subunits of the epithelial Na+ channel (ENaC)
undergo proteolytic processing by furin. Cleavage of the ?-subunit by furin at
the consensus site ?RKRR143 and subsequent cleavage by a second protease at a
distal site strongly activate the channel. For example, coexpression of prostasin
with ENaC increases both channel function and cleavage at the ?RKRK186 site. We
generated a polyclonal antibody that recognizes the region 144-186 in the
?-subunit (anti-?43) to determine whether prostasin promotes the release of the
intervening tract between the putative furin and ?RKRK186 cleavage sites.
Anti-?43 precipitated both full-length (93 kDa) and furin-processed (83 kDa)
?-subunits from extracts obtained from oocytes expressing ??HA-?-V5 channels, but
only the full-length (93 kDa) ?-subunit from oocytes expressing ??HA-?-V5
channels and either wild-type or a catalytically inactive prostasin. Although
both wild-type and catalytically inactive prostasin activated ENaCs in an
aprotinin-sensitive manner, only wild-type prostasin bound to aprotinin beads,
suggesting that catalytically inactive prostasin facilitates the cleavage of the
?-subunit by an endogenous protease in Xenopus oocytes. As dietary salt
restriction increases cleavage of the renal ?-subunit, we assessed release of the
43-mer inhibitory tract on rats fed a low-Na+ diet. We found that a low-Na+ diet
increased ?-subunit cleavage detected with the anti-? antibody and dramatically
reduced the fraction precipitated with the anti-?43 antibody. Our results suggest
that the inhibitory tract dissociates from the ?-subunit in kidneys from rats on
a low-Na+ diet.
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