TY  - JOUR
A1  - Franzen, Rochus
A1  - Fabbro, Doriano
A1  - Aschrafi, Armaz
A1  - Pfeilschifter, Josef
A1  - Huwiler, Andrea
T1  - Nitric oxide induces degradation of the neutral ceramidase in rat renal mesangial cells and is counterregulated by protein kinase C
T2  - Journal of biological chemistry
N2  - Ceramide levels are strongly increased by stimulation of renal mesangial cells with nitric oxide (NO). This effect was shown previously to be due to a dual action of NO, comprising an activation of sphingomyelinases and an inhibition of ceramidase activity. In this study we show that the NO-triggered inhibition of neutral ceramidase activity is paralleled by a down-regulation at the protein level. A complete loss of neutral ceramidase protein is obtained after 24 h of stimulation. Whereas the selective proteasome inhibitor lactacystin blocked NO-evoked ceramidase degradation, several caspase inhibitors were ineffective. Moreover, the NO-induced degradation is reversed by the protein kinase C (PKC) activator, 12-O-tetradecanoylphorbol-13-acetate (TPA), and also by the physiological PKC activators platelet-derived growth factor-BB (PDGF), angiotensin II and ATP, resulting in a normalization of neutral ceramidase protein as well as activity. In vivophosphorylation studies using 32Pi-labeled mesangial cells revealed that TPA, PDGF, angiotensin II, and ATP trigger an increased phosphorylation of the neutral ceramidase, which is blocked by the broad spectrum PKC inhibitor Ro-31 8220 but not by CGP 41251, which has a preferential action on Ca2+-dependent isoforms, thus suggesting the involvement of a Ca2+-independent PKC isoform. In vitro phosphorylation assays using recombinant PKC isoenzymes and neutral ceramidase immunoprecipitated from unstimulated mesangial cells show that particularly the PKC-δ isoform and to a lesser extent the PKC-α isoform are efficient in directly phosphorylating neutral ceramidase. In summary, our data show that NO is able to induce degradation of neutral ceramidase, thereby promoting accumulation of ceramide in the cell. This effect is reversed by PKC activation, most probably by the PKC-δ isoenzyme, which can directly phosphorylate and thereby prevent neutral ceramidase degradation. These novel regulatory interactions will provide therapeutically valuable information to target neutral ceramidase stability and subsequent ceramide accumulation.
Y1  - 2021
UR  - http://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/76001
UR  - https://nbn-resolving.org/urn:nbn:de:hebis:30:3-760013
SN  - 0021-9258
VL  - 277.2002
IS  - 48
SP  - 46184
EP  - 46190
PB  - American Society for Biochemistry and Molecular Biology Publications
CY  - Bethesda, Md
ER  -