Apoptosis and JNK activation are differentially regulated by Fas expression level in renal tubular epithelial cells.
BACKGROUND
In chronic renal disease, renal tubular epithelial cell (RTC) Fas expression is up-regulated, leading to apoptotic RTC deletion and tubular atrophy. In vitro, cytokine- or hypoxia-induced up-regulation of Fas expression is associated with RTC apoptosis. In contrast, constitutively expressed, low level RTC Fas does not mediate apoptosis, suggesting that Fas may be coupled to expression level-dependent RTC signaling pathways. Fas is known to signal through JNK in many systems, but the requirement of JNK activation for apoptosis remains controversial.


METHODS
To determine if RTC Fas regulates JNK activity and apoptosis, human RTC were transfected with graded concentrations of a eukaryotic expression vector for murine Fas. Apoptosis was measured by annexin V, TUNEL and PARP cleavage assays. JNK activity was determined by immune complex kinase assay and/or immunoblots with phospho-specific JNK antibodies, in the presence or absence of co-expressed dominant negative JNK constructs.


RESULTS
Fas antibody stimulation of RTC with high Fas expression levels (to model RTC phenotype in renal disease) caused a tenfold increase in apoptosis, while RTC with low level Fas expression (to model normal RTC phenotype) were apoptosis-resistant. Fas ligation activated JNK in RTC expressing low levels of Fas, but not in apoptosis-sensitive RTC with increased Fas expression. Dominant negative JNK co-expression failed to inhibit apoptosis in RTC expressing high levels of Fas, suggesting that JNK is neither necessary, nor sufficient, for Fas-dependent apoptosis.


CONCLUSIONS
At high levels of expression, RTC Fas promotes apoptosis in a JNK-independent manner. At low basal expression, Fas induces JNK activation, but not apoptosis, consistent with novel roles for RTC Fas as a mediator of cell stress or chronic inflammation.
Khan
S
S
Department of Medicine, Case Western Reserve University School of Medicine, and Rammelkamp Center for Research, MetroHealth Medical Center, Cleveland, Ohio, USA.
Koepke
A
A
Jarad
G
G
Schlessman
K
K
Cleveland
R P
RP
Wang
B
B
Konieczkowski
M
M
Schelling
J R
JR
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000502:physiology; D004789:Enzyme Activation / Q000502:physiology; D004847:Epithelial Cells / Q000502:physiology; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D007687:Kidney Tubules, Proximal / Q000502:physiology*; D051379:Mice; D048055:Mitogen-Activated Protein Kinase 8; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism* / Q000502:physiology; D010957:Plasmids / Q000502:physiology; D014162:Transfection; D019014:fas Receptor / Q000235:genetics / Q000378:metabolism*
D018076:DNA, Complementary; D019014:fas Receptor; D048031:JNK Mitogen-Activated Protein Kinases; D048055:Mitogen-Activated Protein Kinase 8; D020928:Mitogen-Activated Protein Kinases