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. 2016 Mar 2;7(16):21786–21798. doi: 10.18632/oncotarget.7846

Figure 3. Loss of Pin1 does not affect Myc protein stability in B cells and MEFs.

Figure 3

A. c-myc mRNA levels normalized to the TBP mRNA (TATA Box Binding Protein) in control, pre-tumoral splenic B cells and Eμ-myc tumors of the reported Pin1 genotype, as assessed by RT-qPCR. B. Immunoblot analysis of total (Myc), Ser 62-phosphorylated Myc (S62P-Myc) and Thr 58-phosphorylated Myc (T58P-Myc) in B220+ cells purified from spleens of the indicated genotype. Vinculin is shown as loading control. Relative densitometric analysis was performed using the Image Lab 5.0 software. The levels of total Myc normalized to Vinculin, and of S62P-Myc and T58P-Myc normalized to total Myc are reported on the right. In A., B. P-values were calculated using Student's t-test. C. Immunoblot analysis of total Myc and S62P-Myc in Eμ-myc tumors of the reported Pin1 genotype. D. Immunoblot analysis of endogenous Myc in quiescent MEFs (0h) or the same cells stimulated with 20% serum for the indicated times. Relative quantifications, performed as described in B, are reported at the bottom, for either total Myc (left) or S62P-Myc (right). E. Stability of endogenous Myc in asynchronous proliferating Pin1+/+ and Pin1−/− MEFs. Lysates were prepared at the indicated times after treatment of the cells with cycloheximide (CHX) and immunoblotted for total Myc and vinculin. The quantifications of the immunoblot and calculated Myc half-lives are shown at the bottom. Two independent MEF populations of each genotype were analyzed. One representative experiment is shown.