beta-catenin signaling and regulation of cyclin D1 promoter in NRK-49F cells transformed by down-regulation of the tumor suppressor lysyl oxidase.
Lysyl oxidase is the enzyme that is essential for collagen and elastin cross-linking. Previous investigations showed that lysyl oxidase is down-regulated in many human tumors and ras-transformed cells. Recently, we proved that antisense down-regulation of lysyl oxidase in NRK-49F cells induced phenotypic changes and oncogenic transformation, characterized by p21(ras) activation and beta-catenin/cyclin D1 up-regulation. In the present paper, we examined beta-catenin intracellular distribution and its association with E-cadherin. We observed an increased association between E-cadherin and beta-catenin in the lysyl-oxidase down-regulated cells during serum starvation. Moreover, we found that beta-catenin cytoplasmic and nuclear levels were increased, suggesting a failure of its down-regulation by the APC-GSK-3beta system, in particular the GSK-3beta phosphorylation of ser-33/37 and thr-41 of beta-catenin. Finally, we investigated the mechanisms leading to the observed cyclin D1 up-regulation. We showed that in the antisense lysyl oxidase cells the cyclin D1 promoter was activated through the LEF and the ATF/CRE sites in the proximal promoter. While the promoter activation through LEF is compatible with beta-catenin signaling, we investigated the possibility that the CRE-dependent activation might be linked to the down-regulation of lysyl oxidase. In fact, up-regulation of lysyl oxidase in a COS-7 cell model showed a significant diminution of the CREB protein binding to the cyclin D1 promoter, leading to a dramatic inhibition of its activity and a significant down-regulation of cyclin D1 protein level in vivo. Finally, our study describes some major anomalies occurring in lysyl oxidase down-regulated fibroblasts, related to beta-catenin signaling and cyclin D1 expression.
Giampuzzi
Monia
M
Laboratorio di Nefrologia, Istituto G. Gaslini, Largo G. Gaslini, 5, 16147 Genova, Italy.
Oleggini
Roberta
R
Albanese
Chris
C
Pestell
Richard
R
Di Donato
Armando
A
D000818:Animals; D019556:COS Cells; D050882:CREB-Binding Protein / Q000378:metabolism; D015820:Cadherins / Q000378:metabolism; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic / Q000378:metabolism*; D002522:Chlorocebus aethiops; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D005347:Fibroblasts; D015972:Gene Expression Regulation, Neoplastic; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D008249:Protein-Lysine 6-Oxidase / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*; D051176:beta Catenin / Q000378:metabolism*
D015820:Cadherins; D051176:beta Catenin; D019938:Cyclin D1; D008249:Protein-Lysine 6-Oxidase; D050882:CREB-Binding Protein; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D038362:Glycogen Synthase Kinase 3