Figure 2
Increased oxaliplatin resistance in SW480 cells expressing the TAP-YB-1 construct and validation of TAP-YB-1 interaction with six identified protein from the LC-MS/MS. A) Representative Western blot analysis of SW480 clones expressing the TAP or the TAP-YB-1 constructs. β-actin was used as loading controls. The histogram below the Western blots indicates the endogenous and TAP-YB-1 expression levels in the indicated clones. B) Survival assays of SW480 clones expressing TAP or TAP-YB-1 constructs. Cells were treated with one or two μM of oxaliplatin for 18 h. Live cells were then counted with a hemacytometer by the trypan blue exclusion technique. Experiments were performed in duplicate. Bars represent SEM. C) Clonogenic assays with SW480 clones expressing TAP or TAP-YB-1 constructs. Cells were treated with the indicated concentrations of oxaliplatin for 18 hours and then plated in 90-mm petri dishes with fresh media without drugs for 10 days. Survival fraction represents the number of colonies after treatment divided by the number of colonies without treatment (in %). Results represent the mean ± SEM of duplicate experiments. D) TAP-YB-1 interaction of HNRPA2B1, HNRPL, MRPL13, NONO, RALY, and RIBC2 with the YB-1 protein. Proteins from TAP and TAP-YB-1 expressing SW480 cells were eluted from the streptavidin beads and analyzed by SDS-PAGE with antibodies against the indicated proteins. Proteins were revealed with an ECL Plus kit. WCE represents the whole cell lysate. TAP represents the protein eluted from the streptavidin beads.