Role of PKCα in feedback regulation of Na⁺transport in an electrically tight epithelium

Abstract

It has long been known that Na+ channels in electrically tight epithelia are regulated by homeostatic mechanisms that maintain a steady state and allow new levels of transport to be sustained in hormonally challenged cells. Little is known about the potential pathways involved in these processes. In addition to short-term effect, recent evidence also indicates the involvement of PKC in the long-term regulation of the epithelial Na+ channel (ENaC) at the protein level (40). To determine whether stimulation of ENaC involves feedback regulation of PKC levels, we utilized Western blot analysis to determine the distribution of PKC isoforms in polarized A6 epithelia. We found the presence of PKC isoforms in the conventional (α and γ), novel (δ, η, and ε), and atypical (ι, λ, and ζ) groups. Steady-state stimulation of Na+ transport with aldosterone was accompanied by a specific decrease of PKCα protein levels in both the cytoplasmic and membrane fractions. Similarly, overnight treatment with an uncharged amiloride analog (CDPC), a procedure that through feedback regulation causes a stimulation of Na+ transport, also decreased PKCα levels. These effects were additive, indicating separate mechanisms that converge at the level of PKCα. These effects were not accompanied by changes of PKCα mRNA levels as determined by Northern blot analysis. We propose that this may represent a novel regulatory feedback mechanism necessary for sustaining an increase of Na+ transport.

The A‐kinase anchoring protein 15 regulates feedback inhibition of the epithelial Na+channel

“…As shown in Fig. 9A, coexpression with this kinase isoform reduced ENaC activity to ϳ28% of baseline, consistent with the premise that conventional PKC isoforms inhibit ENaC (11). In the presence of AKAP15m, this inhibition is attenuated to 72% of baseline, indicating that channel inhibition by PKC␣ and presumably feedback inhibition can indeed These data indicate that ENaC surface density is unchanged between ENaC (gray) and ENaCϩAKAP15 (black) coexpressing oocytes.…”

“…A potential candidate for this interaction would be PKC, which is involved in the feedback inhibition of ENaC by elevated [Na ϩ ] i . More specifically, PKC␣ was previously implicated in the feedback regulation of ENaC (11). This isoform is endogenously expressed in oocytes; it is inhibitory of ENaC (see below and ref.…”

“…The PKC isoforms implicated in this response were not examined. However, we subsequently established a specific function for PKC␣ in the process of feedback regulation of ENaC (11). We also proposed that this isoform may play a role in feedback inhibition by intracellular Na ϩ .…”

A long isoform of the epithelial sodium channel alpha subunit forms a highly active channel

“…Two types of inhibition by sodium have been described and differentiated based on time course: 12-14 a slow effect due to high [Na + ], termed "feedback inhibition," and mediated by PKC 15 and a fast effect termed "self-inhibition" likely mediated by the Na + ions interaction with the channel, 11 and this represents an intrinsic channel property that does not require further second messenger. 14 ENaC in native epithelia, and especially in the kidney, is exposed to different [Na + ] making both regulatory processes physiologically relevant.…”

Brakes and gas-regulation of ENaC by sodium

“…This has been recognized early on and termed feedback inhibition. 4 Feedback inhibition is ubiquitously observed for ENaC in many native and heterologous systems and has been reported to involve the ubiquitin ligase protein Nedd4-2 5 , and protein kinase C. 6 The physiological significance of feedback inhibition has been demonstrated by Palmer and colleagues 7 ; however, an exact link to the channel has remained missing.…”