Wnt4 induces nephronic tubules in metanephric mesenchyme by a non-canonical mechanism.
Wnt4 and β-catenin are both required for nephrogenesis, but studies using TCF-reporter mice suggest that canonical Wnt signaling is not activated in metanephric mesenchyme (MM) during its conversion to the epithelia of the nephron. To better define the role of Wnt signaling, we treated rat metanephric mesenchymal progenitors directly with recombinant Wnt proteins. These studies revealed that Wnt4 protein, which is required for nephron formation, induces tubule formation and differentiation markers Lim1 and E-cadherin in MM cells, but does not activate a TCF reporter or up regulate expression of canonical Wnt target gene Axin-2 and has little effect on the stabilization of β-catenin or phosphorylation of disheveled-2. Furthermore, Wnt4 causes membrane localization of ZO-1 and occludin in tight junctions. To directly examine the role of β-catenin/TCF-dependent transcription, we developed synthetic cell-permeable analogs of β-catenin's helix C, which is required for transcriptional activation, in efforts to specifically inhibit canonical Wnt signaling. One inhibitor blocked TCF-dependent transcription and induced degradation of β-catenin but did not affect tubule formation and stimulated the expression of Lim1 and E-cadherin. Since a canonical mechanism appears not to be operative in tubule formation, we assessed the involvement of the non-canonical Ca(2+)-dependent pathway. Treatment of MM cells with Wnt4 induced an influx of Ca(2+) and caused phosphorylation of CaMKII. Moreover, Ionomycin, a Ca(2+)-dependent pathway activator, stimulated tubule formation. These results demonstrate that the canonical Wnt pathway is not responsible for mesenchymal-epithelial transition (MET) in nephron formation and suggest that the non-canonical calcium/Wnt pathway mediates Wnt4-induced tubulogenesis in the kidney.
Cancer and Developmental Biology Laboratory, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702-1201, USA.
Shunsuke
S
Tanigawa
Honghe
H
Wang
Yili
Y
Yang
Nirmala
N
Sharma
Nadya
N
Tarasova
Rieko
R
Ajima
Terry P
TP
Yamaguchi
Luis G
LG
Rodriguez
Alan O
AO
Perantoni
D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007684:Kidney Tubules / Q000166:cytology / Q000187:drug effects / Q000196:embryology / Q000378:metabolism; D008648:Mesoderm / Q000166:cytology / Q000187:drug effects* / Q000196:embryology* / Q000378:metabolism; D051379:Mice; D008954:Models, Biological; D009024:Morphogenesis / Q000187:drug effects; D009399:Nephrons / Q000166:cytology / Q000187:drug effects* / Q000196:embryology* / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D051820:TCF Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects / Q000235:genetics; D051153:Wnt Proteins / Q000494:pharmacology*; D060528:Wnt4 Protein; D051176:beta Catenin / Q000737:chemistry / Q000378:metabolism
D051820:TCF Transcription Factors; C555581:WNT4 protein, human; D051153:Wnt Proteins; D060528:Wnt4 Protein; C555582:Wnt4 protein, mouse; C555583:Wnt4 protein, rat; D051176:beta Catenin; D015759:Ionomycin; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2
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