Transcription Factor AP-2α Promotes EGF-Dependent Invasion of Human Trophoblast

The basic helix-span-helix transcription factor activating protein (AP)-2 alpha 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, we here show that AP-2 alpha 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-2 alpha 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-2 alpha small interfering RNA decreased EGF-dependent invasion. Proliferation of SGHPL-5 cells and primary EVTs, measured by cumulative cell numbers and 5-bromo-2'-deoxyuridine labeling, respectively, were not affected on loss of AP-2 alpha. EGF-dependent induction of matrix metalloproteinase (MMP)-2, pro- and active form of urokinase plasminogen activator, and chorionic gonadotropin (CG)-beta was noticed in shRNAmir-control cells, whereas these genes were suppressed in EGF-treated shRNAmir-AP-2 alpha cells. Similarly, EGF-stimulated MMP-2 and CG beta protein expression was reduced in AP-2 alpha small interfering RNA-treated primary EVTs. Knockdown of AP-2 alpha also decreased luciferase activity of the CG beta 5 promoter in SGHPL-5 cells, which was compensated upon transient overexpression of AP-2 alpha cDNA. In conclusion, we show that AP-2 alpha 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. (Endocrinology 152: 1458-1469, 2011)