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Mammalian Nemo-like kinase enhances β-catenin-TCF transcription activity in human osteosarcoma and neuroblastoma cells.

Yasuda J, Ichikawa H - Proc. Jpn. Acad., Ser. B, Phys. Biol. Sci. (2007)

Bottom Line: The nuclear localization of Lymphoid enhancer factor 1 (LEF1) and β-catenin ΔN was not altered by NLK overexpression regardless of its effect on β-catenin-TCF complex activity.Mutations in the potential NLK phosphorylation sites in β-catenin did not change its transcription activity either.Our results suggest that NLK positively regulates Wnt/β-catenin signaling in a cell type dependent manner through an unidentified mechanism.

View Article: PubMed Central - PubMed

Affiliation: Cancer Transcriptome Project, National Cancer Center Research Institute, Tokyo, Japan .

ABSTRACT
Nemo-like kinase (NLK) is an evolutionarily conserved serine/threonine kinase and has been considered to be a suppressor of Wnt signaling in mammalian cells. Our study, however, has raised the possibility that NLK also functions as a Wnt signaling activator. In human osteosarcoma and neuroblastoma cell lines, NLK specifically enhanced β-catenin-TCF complex transcription activity. The effect required kinase activity of NLK and co-expression of the β-catenin ΔN (constitutive active mutant of β-catenin). The nuclear localization of Lymphoid enhancer factor 1 (LEF1) and β-catenin ΔN was not altered by NLK overexpression regardless of its effect on β-catenin-TCF complex activity. Reporter analysis using LEF1 mutants at known NLK target sites indicated that NLK may have different activation targets for β-catenin-TCF complex. Mutations in the potential NLK phosphorylation sites in β-catenin did not change its transcription activity either. Our results suggest that NLK positively regulates Wnt/β-catenin signaling in a cell type dependent manner through an unidentified mechanism.

No MeSH data available.


Related in: MedlinePlus

Overexpression of NLK-S did not change the nuclear localization of LEF1 and stabilized β-catenin in 293 cells (A) or SaOS-2 cells (B). Immunofluorescence analysis of LEF1 and β-catenin without (upper row) and with (lower row) NLK-S. For both cell lines, 1×105 of cells were transfected with 1 μg each of pcDNA3-HA-LEF1, pcDNA3-MYC-β-catenin ΔN with or without 1 μg pcDNA3-NLK-S in 6-well plate wells.
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f4-83_016: Overexpression of NLK-S did not change the nuclear localization of LEF1 and stabilized β-catenin in 293 cells (A) or SaOS-2 cells (B). Immunofluorescence analysis of LEF1 and β-catenin without (upper row) and with (lower row) NLK-S. For both cell lines, 1×105 of cells were transfected with 1 μg each of pcDNA3-HA-LEF1, pcDNA3-MYC-β-catenin ΔN with or without 1 μg pcDNA3-NLK-S in 6-well plate wells.

Mentions: It is known that the transcription activity of the worm TCF homolog POP-1 is regulated through its subcellular localization.14), 15), 35), 36) When the Wnt signaling is activated, the nuclear concentration of POP-1 is reduced and remaining nuclear POP-1 binds to β-catenin homologs and mediates the signal.35), 36) If the same mechanism in C. elegans occurred in some mammalian cells such as SaOS-2, NLK would reduce the nuclear localization of the TCF/LEF transcription factors for activation of Wnt signaling. It is an interesting question if NLK could differently regulate the localization of TCF between 293 cells and SaOS-2 cells. On the other hand, β-catenin ΔN usually accumulates in the nucleus to activate target genes. To examine whether subcellular localization of β-catenin ΔN is differently regulated by NLK depending on the cell type, Hemagglutinin (HA) tagged LEF1 (a member of the TCF transcription factor family) and β-catenin ΔN were co-expressed with NLK in either 293 or SaOS-2 cells. Co-expression with NLK did not alter the nuclear localization of either LEF1 or β-catenin ΔN in either 293 or SaOS-2 cells (Fig. 4A and B, respectively).


Mammalian Nemo-like kinase enhances β-catenin-TCF transcription activity in human osteosarcoma and neuroblastoma cells.

Yasuda J, Ichikawa H - Proc. Jpn. Acad., Ser. B, Phys. Biol. Sci. (2007)

Overexpression of NLK-S did not change the nuclear localization of LEF1 and stabilized β-catenin in 293 cells (A) or SaOS-2 cells (B). Immunofluorescence analysis of LEF1 and β-catenin without (upper row) and with (lower row) NLK-S. For both cell lines, 1×105 of cells were transfected with 1 μg each of pcDNA3-HA-LEF1, pcDNA3-MYC-β-catenin ΔN with or without 1 μg pcDNA3-NLK-S in 6-well plate wells.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3756733&req=5

f4-83_016: Overexpression of NLK-S did not change the nuclear localization of LEF1 and stabilized β-catenin in 293 cells (A) or SaOS-2 cells (B). Immunofluorescence analysis of LEF1 and β-catenin without (upper row) and with (lower row) NLK-S. For both cell lines, 1×105 of cells were transfected with 1 μg each of pcDNA3-HA-LEF1, pcDNA3-MYC-β-catenin ΔN with or without 1 μg pcDNA3-NLK-S in 6-well plate wells.
Mentions: It is known that the transcription activity of the worm TCF homolog POP-1 is regulated through its subcellular localization.14), 15), 35), 36) When the Wnt signaling is activated, the nuclear concentration of POP-1 is reduced and remaining nuclear POP-1 binds to β-catenin homologs and mediates the signal.35), 36) If the same mechanism in C. elegans occurred in some mammalian cells such as SaOS-2, NLK would reduce the nuclear localization of the TCF/LEF transcription factors for activation of Wnt signaling. It is an interesting question if NLK could differently regulate the localization of TCF between 293 cells and SaOS-2 cells. On the other hand, β-catenin ΔN usually accumulates in the nucleus to activate target genes. To examine whether subcellular localization of β-catenin ΔN is differently regulated by NLK depending on the cell type, Hemagglutinin (HA) tagged LEF1 (a member of the TCF transcription factor family) and β-catenin ΔN were co-expressed with NLK in either 293 or SaOS-2 cells. Co-expression with NLK did not alter the nuclear localization of either LEF1 or β-catenin ΔN in either 293 or SaOS-2 cells (Fig. 4A and B, respectively).

Bottom Line: The nuclear localization of Lymphoid enhancer factor 1 (LEF1) and β-catenin ΔN was not altered by NLK overexpression regardless of its effect on β-catenin-TCF complex activity.Mutations in the potential NLK phosphorylation sites in β-catenin did not change its transcription activity either.Our results suggest that NLK positively regulates Wnt/β-catenin signaling in a cell type dependent manner through an unidentified mechanism.

View Article: PubMed Central - PubMed

Affiliation: Cancer Transcriptome Project, National Cancer Center Research Institute, Tokyo, Japan .

ABSTRACT
Nemo-like kinase (NLK) is an evolutionarily conserved serine/threonine kinase and has been considered to be a suppressor of Wnt signaling in mammalian cells. Our study, however, has raised the possibility that NLK also functions as a Wnt signaling activator. In human osteosarcoma and neuroblastoma cell lines, NLK specifically enhanced β-catenin-TCF complex transcription activity. The effect required kinase activity of NLK and co-expression of the β-catenin ΔN (constitutive active mutant of β-catenin). The nuclear localization of Lymphoid enhancer factor 1 (LEF1) and β-catenin ΔN was not altered by NLK overexpression regardless of its effect on β-catenin-TCF complex activity. Reporter analysis using LEF1 mutants at known NLK target sites indicated that NLK may have different activation targets for β-catenin-TCF complex. Mutations in the potential NLK phosphorylation sites in β-catenin did not change its transcription activity either. Our results suggest that NLK positively regulates Wnt/β-catenin signaling in a cell type dependent manner through an unidentified mechanism.

No MeSH data available.


Related in: MedlinePlus