Journal: Oncotarget

Article Title: CRKL mediates EML4-ALK signaling and is a potential therapeutic target for ALK-rearranged lung adenocarcinoma

doi: 10.18632/oncotarget.8638

Figure Lengend Snippet: A. Regulatory network sensitive to ALK inhibitors in H3122 and H2228 cells revealed by phosphotyrosine peptide mapping. Core signaling proteins inhibited by ALK inhibitors in H3122 and H2228 cells are shown in green. The proteins with ≥2-fold decrease of phosphorylation (at least one tyrosine residue) 1 hr after treatment are presented. B. IPA analysis of tyrosine-phosphorylated proteins with differential signaling pathways in H3122 and H2228 cells treated with ALK inhibitor. The yellow line indicates the fraction associated with each pathway of genes that were expressed in each cell line. C. Networking of CRKL with various signaling molecules detected in the phosphotyrosine peptide mapping study. Core signaling molecules inhibited by ALK inhibitors in H3122 and H2228 cells are shown in green. D. Validation of decreased CRKL phosphorylation identified by mass spectrometry. Lysates from H2228 and H3122 cells treated with/without crizotinib (900nM) or NMS-E628 (300nM) for 1 hr were subjected to Western blot probed with anti-p-CRKL (Y207) antibody. As controls, total CRKL and Tubulin were also detected. E. Effect of ALK siRNA knockdown (20 nM for 72 h) on CRKL phosphorylation. Western blot analyses were performed on the lysates from H3122 and H2228 cells treated with ALK siRNAs (four individual siRNA or their smartpool at 20 nM for 72 h) to determine p-CRKL (Y207) level. The graph shows the quantification of p-CRKL levels for each treatment (Student's t -test: p -value < 0.05).

Article Snippet: Anti-ALK, anti-phospho-ALK (Tyr 1604 ), anti-phospho-ALK (Tyr 1096 ), anti-phospho-ALK (Tyr 1078 ), anti-phospho-CRKL (Tyr 207 ), anti-CRKL, anti-ERK1/2 and anti-phospho-ERK1/2 antibodies were obtained from Cell Signaling Technology.

Techniques: Mass Spectrometry, Western Blot

Journal: Oncotarget

Article Title: CRKL mediates EML4-ALK signaling and is a potential therapeutic target for ALK-rearranged lung adenocarcinoma

doi: 10.18632/oncotarget.8638

Figure Lengend Snippet: A. Western blot analysis showing the knockdown of ALK and CRKL in ALK siRNA- and CRKL siRNA-transfected MP038 cells. B. Viability of MP038 cells after transfected with ALK siRNAs or CRKL siRNAs for 72 h. Cell viability (in triplicate) was measured by MTS assay (Student's t -test: p -value < 0.05).

Article Snippet: Anti-ALK, anti-phospho-ALK (Tyr 1604 ), anti-phospho-ALK (Tyr 1096 ), anti-phospho-ALK (Tyr 1078 ), anti-phospho-CRKL (Tyr 207 ), anti-CRKL, anti-ERK1/2 and anti-phospho-ERK1/2 antibodies were obtained from Cell Signaling Technology.

Techniques: Western Blot, Transfection, MTS Assay

Journal: Oncotarget

Article Title: CRKL mediates EML4-ALK signaling and is a potential therapeutic target for ALK-rearranged lung adenocarcinoma

doi: 10.18632/oncotarget.8638

Figure Lengend Snippet: H3122 and H2228 cells were treated with dasatinib (0, 10, 100, 1000 nM) and/or crizotinib (100 nM), and cell lysates were prepared 1 hr after the indicated treatment. Western blot analyses were performed to examine the statuses of CRKL, ALK and ERK tyrosine phosphorylation. Tubulin was used as a loading control.

Article Snippet: Anti-ALK, anti-phospho-ALK (Tyr 1604 ), anti-phospho-ALK (Tyr 1096 ), anti-phospho-ALK (Tyr 1078 ), anti-phospho-CRKL (Tyr 207 ), anti-CRKL, anti-ERK1/2 and anti-phospho-ERK1/2 antibodies were obtained from Cell Signaling Technology.

Techniques: Western Blot