Transcription factor AP-2alpha promotes EGF-dependent invasion of human trophoblast

The basic helix-span-helix transcription factor activating protein (AP)-2I? is critically involved in cell-specific hormone expression of syncytializing human trophoblasts. Its role in invasive trophoblast differentiation, however, remains largely elusive. Using RT-PCR, Western blotting, and immunofluorescence of first-trimester placentae,wehereshowthat AP-2I? is expressed in extravillous trophoblasts (EVTs) both in situ and in vitro as well as in invasive trophoblast cell lines. Its protein expression was increased upon supplementation of epidermal growth factor (EGF) both in primary EVTs and trophoblastic SGHPL-5 cells. Gene silencing of AP-2I? using small hairpin microRNA (shRNAmir) did not affect basal invasion of SGHPL-5 cells through Matrigel-coated filters but reduced EGF-stimulated invasion. Similarly, treatment of primary EVTs with AP-2I? small interfering RNA decreased EGF-dependent invasion. Proliferation of SGHPL-5 cells and primary EVTs, measured by cumulative cell numbers and 5-bromo-2a??-deoxyuridine labeling, respectively, were not affected on loss of AP-2I?. EGF-dependent induction of matrix metalloproteinase (MMP)-2, proand active form of urokinase plasminogen activator, and chorionic gonadotropin (CG)-I? was noticed in shRNAmir-control cells, whereas these genes were suppressed in EGF-treated shRNAmir-AP-2I? cells. Similarly, EGF-stimulated MMP-2 and CGI? protein expression was reduced in AP-2I? small interfering RNA-treated primary EVTs. Knockdown of AP-2I? also decreased luciferase activity of the CGI?5 promoter in SGHPL-5 cells, which was compensated upon transient overexpression of AP-2I? cDNA. In conclusion, we show that AP-2I? expression positively affects human trophoblast invasion under EGF-stimulated conditions, likely by inducing critical invasion-promoting genes such MMP-2, urokinase plasminogen activator, and CG. Copyright A? 2011 by The Endocrine Society.