Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Sequences of the integrin β1, β1A, β1C, hnRNP E1-GFP and β-actin PCR primers.

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques:

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Activity of Fyn regulates the phosphorylation and nuclear localization of hnRNP E1 via PAK1. (A–C) Western blot analysis revealed the relative expression levels of hnRNP E1 and PAK1 protein among BxPC3, BxPC3GFP and BxPC3KdFyn cells. β-actin blotting was used as a loading control. (D and E) Co-immunoprecipitation analysis showed the threonine-phosphorylation of hnRNP E1 and the interaction of hnRNP E1 with PAK1 among BxPC3, BxPC3GFP and BxPC3KdFyn cells. (F and G) Confocal microscopy analysis showed that the nuclear distribution of hnRNP E1 was increased in BxPC3KdFyn cells compared with BxPC3GFP cells. * P<0.05. Data are presented as the mean ± standard deviation from ≥3 independent experiments.

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Activity Assay, Phospho-proteomics, Western Blot, Expressing, Control, Immunoprecipitation, Confocal Microscopy, Standard Deviation

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Fyn regulates the alternative splicing of integrin β1 via hnRNP E1. (A) Co-immunoprecipitation analysis showed the expression of GFP-hnRNP E1 fusion protein in BxPC3 cells. (B, F and G) Splicing of integrin β1 was analyzed under different Fyn activity levels or hnRNP E1 expression levels in BxPC3 cells. (C–E) Knockdown of hnRNP E1 expression decreased the phosphorylation of hnRNP E1 in BxPC3 cells. (H and I) Splicing of integrin β1 was analyzed under different Fyn activity levels or hnRNP E1 expression levels in Panc1 cells. (J) RNA immunoprecipitation showed the interaction between hnRNP E1 and the pre-mRNA of integrin β1. * P<0.05. Data are presented as the mean ± standard deviation from ≥3 independent experiments. Sequences of the PCR primers for integrin β1 pre-mRNA, which cover the exon and intron sequences of integrin β1C are listed in .

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Alternative Splicing, Immunoprecipitation, Expressing, Activity Assay, Knockdown, Phospho-proteomics, RNA Immunoprecipitation, Standard Deviation

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Proteins identified by mass spectrometry in the hnRNP E1 spliceosome of BxPC3KdFyn cells.

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Mass Spectrometry, Protein Binding

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: hnRNP E1 spliceosome regulates integrin β1 splicing by hnRNP A1 and SF2/ASF. (A) Schematic representation of the genomic sequence of integrin β1. Exon sequences are shown as boxes and introns as solid lines. The splicing pattern used to generate β1A and C transcripts are depicted, and the potential SR protein-binding site is indicated with a vertical arrow. The positions of the stop codons for the various splice variants are also indicated by arrows. The horizontal arrows indicate the primers used in PCR experiments . (B–D) Western blot analysis and co-immunoprecipitation were used to analyze the expression levels and abundance of hnRNP E1-specific spliceosome components among BxPC3, BxPC3GFP and BxPC3KdFyn cells. β-actin blotting was used as a loading control. (E and F) Co-immunoprecipitation and splicing analysis were used to explore the functions of hnRNP A1 and SF2/ASF on the construction of the hnRNP E1 spliceosome complex and the splicing of integrin β1. (G) RNA immunoprecipitation showed that hnRNP A1 and SF2/ASF, but not hnRNP E1, were directly bound to the pre-mRNA of integrin β1. * P<0.05. Data are presented as the mean ± standard deviation from ≥3 independent experiments.

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Sequencing, Protein Binding, Western Blot, Immunoprecipitation, Expressing, Control, RNA Immunoprecipitation, Standard Deviation

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Effect of Fyn and hnRNP E1 on the invasion and metastasis of pancreatic cancer cells. (A–C) Transwell and wound-healing assays revealed that the activity of Fyn and/or the expression of hnRNP E1 significantly affected the invasion and migration abilities of BxPC3 pancreatic cancer cells in vitro . (D–F) An assay of the subcutaneous tumor mass and liver metastatic nodules revealed that the activity of Fyn and/or the expression of hnRNP E1 significantly affected the tumorigenesis and metastasis of BxPC3 pancreatic cancer cells in vivo . Each group of xenograft experiments included 5 mice. * P<0.05. Data are presented as the mean ± standard deviation from ≥3 independent experiments.

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Activity Assay, Expressing, Migration, In Vitro, In Vivo, Standard Deviation

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Associations between the expression of integrin β1A, hnRNP E1 and the categorical clinicopathological parameters of pancreatic carcinoma.

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Expressing

Journal: International Journal of Oncology

Article Title: Fyn/heterogeneous nuclear ribonucleoprotein E1 signaling regulates pancreatic cancer metastasis by affecting the alternative splicing of integrin β1

doi: 10.3892/ijo.2017.4018

Figure Lengend Snippet: Correlation between the expressions of β1A (mRNA) and hnRNP E1 (protein).

Article Snippet: After blocking with 10% goat serum for 30 min, the sections were incubated overnight at 4°C with the hnRNP E1 rabbit polyclonal antibody (sc-28725, 1:200; Santa Cruz Biotechnology).

Techniques: Expressing