Ischemia-induced cleavage of cadherins in NRK cells is not sufficient for beta-catenin transcriptional activity.
Although ischemia is associated with disruption of cadherin-mediated adhesion in renal cell lines, the impact of decreased cadherin function on the transcriptional activity of beta-catenin remains poorly defined. In these studies, we used a simulated ischemia model in normal rat kidney (NRK) cells to disrupt cadherin function. Cell viability; cadherin/catenin expression, function, and localization; and beta-catenin-mediated transcriptional activity were assessed during ischemia/reperfusion. Following 6 hr of ischemia, a decrease in the expression of E- and N-cadherin was seen that correlated with altered cell morphology indicative of decreased intercellular adhesion. While ischemia was associated with activation of glycogen synthase kinase 3 beta (GSK-3beta), this did not correlate with increased phosphorylation of beta-catenin as assessed by Western blots using phosphoryl-specific antibodies. beta-Catenin was not localized to the nucleus by immunofluorescence in ischemic NRK cells, but rather a strong perinuclear signal was seen in reperfused cells. This was consistent with the finding that neither ischemia nor reperfusion activated the transcriptional activity of beta-catenin as assessed by the TCF-optimal promoter (TOPFlash) construct. However, NRK cells possess a competent Wnt pathway, as challenge with lithium chloride elicited a ten-fold increase in luciferase activity. These results suggest that ischemia-induced disruption of cadherin/catenin complexes is not sufficient to stimulate beta-catenin transcriptional activity in NRK cells.
Chen
Gang
G
Department of Systems Biology and Translational Medicine, College of Medicine, Texas A&M University System Health Science Center, Texas, USA.
Akintola
Adebayo D
AD
Catania
Jeffrey M
JM
Covington
Marisa D
MD
Dean
Dana D
DD
Trzeciakowski
Jerome P
JP
Burghardt
Robert C
RC
Parrish
Alan R
AR
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism*; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D047468:Immunoprecipitation; D007511:Ischemia / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008156:Luciferases / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D015398:Signal Transduction; D051820:TCF Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D051153:Wnt Proteins / Q000378:metabolism; D051176:beta Catenin / Q000235:genetics* / Q000378:metabolism
D015820:Cadherins; D051820:TCF Transcription Factors; D051153:Wnt Proteins; D051176:beta Catenin; D000255:Adenosine Triphosphate; D008156:Luciferases; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D038362:Glycogen Synthase Kinase 3