Figure 5. PTPN12 Is Regulated by the REST Tumor Suppressor via miR-124.

(A) Loss of REST expression in human breast cancer. Primary human breast cancers (n = 185) were analyzed by immunohistochemistry for REST expression. Representative panels exhibiting negative and positive REST expression in invasive breast cancers.

(B) Loss of REST expression strongly correlates with loss of PTPN12 expression. Primary human breast cancers (n = 185) were analyzed by immunohisto-chemistry for REST and PTPN12 expression. The level of PTPN12 (y axis) is plotted for tumors with absent, intermediate, or high REST levels (x axis). The median and mean PTPN12 values for each group are represented by a solid red line and plus symbol, respectively. The boxes represent the 25th to 75th percentiles. Association between PTPN12 and REST expression was tested by Fisher’s exact test. Error bars represent maximum and minimum observations within inner fences.

(C) Ectopic REST expression increases PTPN12 protein levels in REST-deficient TNBC cells. HCC70 TNBC cells were transduced with control or REST cDNA, cultured for 9 days, and analyzed for expression of REST and PTPN12 by western.

(D) Model for REST regulation of PTPN12 expression. REST regulates transcription of the neuronal microRNA miR-124. The PTPN12 3′UTR contains three conserved binding sites for miR-124. The sequences surrounding the three miR-124 binding sites are shown for human and five other vertebrate species.

(E) Ectopic miR-124 expression decreases PTPN12 protein levels in HMECs. HMECs were transduced with control or miR-124-containing plasmid, cultured for 7 days, and analyzed for PTPN12 expression by western.

(F) Ectopic miR-124 expression transforms TLM-HMECs. Cells from (E) were assessed for anchorage-independent proliferation.

Error bars represent standard error.