PORCN regulates a subset of genes in a Wnt-independent manner.

<p>A. Gene expression analysis of MDA-MB-231 cells 48 hours after mock transfected (NT) or transfected with 100 nM of indicated siRNA for 48 hrs. PORCN and WLS differentially regulate a distinct subset of genes. B. To illustrate genes differentially regulated by PORCN, the abundance of the indicated transcripts were assessed by qRT-PCR 48 hours after transfection of MDA-MB-231 cells with the indicated siRNAs C. Two independent PORCN siRNAs cause decrease in APEH protein. Immunoblot analysis of APEH and Actin from MDA-MB-231 cells treated with 100 nM Control (C), PORCN7 (P7) or PORCN8 (P8) siRNA for 72 hrs. D. PORCN knockdown triggers loss of APEH in multiple cell lines. Immunoblot analysis of APEH from MCF7, T47D, and DLD-1 cells transfected with 100 nM Control (C), or PORCN7 (P7) siRNAs for 72 hrs. Top panel is 10 s exposure, middle panel is 30 s exposure. Actin (bottom panel) serves as a load control. E. Loss of APEH is blocked by RNAi-immune wildtype and mutant PORCN. Immunoblot analysis of APEH and HA-tagged PORCN in MDA-MB-231 cells co-transfected with P7-resistant WT or H341A PORCN and 100 nM Control (C) or PORCN7 (P7) siRNA for 48 hrs. * = non-specific band recognized by anti-HA antibody, used as load control. Experiment done in duplicate with same results. F. Model of PORCN's function in transformed epithelial cells. PORCN is involved in the acylation and secretion of Wnt proteins. Independent of its role in Wnt secretion, PORCN regulates proliferation and gene expression. The role in gene expression may lead to changes in proliferation.