Association of TCTP with Pim-3 in human pancreatic cancer cell lines.

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Association of TCTP with Pim-3 in human pancreatic cancer cell lines. Association of TCTP with Pim-3 in human pancreatic cancer cell lines. A, co-immunoprecipitation of TCTP and Pim-3 in transiently transfected HEK293T cells. Cell lysates were obtained from HEK293T cells transiently transfected with pcDNA4-Pim-3 and pcDNA4-TCTP-His. The resultant lysates were subjected to immunoprecipitation and immunoblotting. B, co-immunoprecipitation of endogenous TCTP and Pim-3 in PCI55 cells. Cell lysates were obtained from PCI55 cells. The resultant lysates were subjected to immunoprecipitation and immunoblotting. One tenth of the cell lysates from PCI55 were subjected to immunoblotting with anti-TCTP or anti-Pim-3 antibodies as positive controls. C, colocalization of TCTP and Pim-3 in human pancreatic cancer cells. PCI55 cells were immunostained with a combination of anti-TCTP and anti-Pim-3 antibodies as described in Materials and Methods. The fluorescent images were digitally merged. Yellow coloration in overlay panels indicates colocalization of Pim-3 and TCTP. Nuclei were counterstained with 4′,6-diamidino-2-phenylindole. D, TCTP and Pim-3 expression in human pancreatic cancer cell lines was detected by Western blotting. β-Actin was used as an internal control. E, the expression levels of TCTP, Pim-3, and β-actin were quantified using NIH ImageJ software. TCTP/β-actin and Pim-3/β-actin ratios were calculated for each cell line. F, correlation between TCTP and Pim-3 expression in human pancreatic cancer cell lines, with a linear regression line and Spearman correlation significance (Spearman correlation coefficient = 1, P < 0.01). G, schematic representation of cDNA constructs for each TCTP deletion mutant. H, cell lysates were obtained from HEK293T cells transiently cotransfected with Pim-3 and TCTP deletion mutants. The resultant lysates were immunoprecipitated and immunoblotted. I, schematic representation of cDNA constructs for each Pim-3 deletion mutant. J, cell lysates were obtained from HEK293T cells transiently cotransfected with TCTP and Pim-3 deletion mutants. The resultant lysates were immunoprecipitated and immunoblotted. K, schematic representation of the presumed interaction between Pim-3 and TCTP proteins. Fei Zhang et al. Mol Cancer Res 2013;11:1508-1520 ©2013 by American Association for Cancer Research