Krüppel-Like Factor 6 Expression Changes during Trophoblast Syncytialization and Transactivates ßhCG and PSG Placental Genes

BACKGROUND: Krüppel-like factor-6 (KLF6) is a widely expressed member of the Sp1/KLF family of transcriptional regulators involved in differentiation, cell cycle control and proliferation in several cell systems. Even though the highest expression level of KLF6 has been detected in human and mice placenta, its function in trophoblast physiology is still unknown. METHODOLOGY/PRINCIPAL FINDINGS: Herein, we explored KLF6 expression and sub-cellular distribution in human trophoblast cells differentiating into the syncytial pathway, and its role in the regulation of genes associated with placental development and pregnancy maintenance. Confocal immunofluorescence microscopy demonstrated that KLF6 is expressed throughout human cytotrophoblast differentiation showing no evident modifications in its nuclear and cytoplasmic localization pattern. KLF6 transcript and protein peaked early during the syncytialization process as determined by qRT-PCR and western blot assays. Overexpression of KLF6 in trophoblast-derived JEG-3 cells showed a preferential nuclear signal correlating with enhanced expression of human β-chorionic gonadotropin (βhCG) and pregnancy-specific glycoprotein (PSG) genes. Moreover, KLF6 transactivated βhCG5, PSG5 and PSG3 gene promoters. Deletion of KLF6 Zn-finger DNA binding domain or mutation of the consensus KLF6 binding site abolished transactivation of the PSG5 promoter. CONCLUSIONS/SIGNIFICANCE: Results are consistent with KLF6 playing a role as transcriptional regulator of relevant genes for placental differentiation and physiology such as βhCG and PSG, in agreement with an early and transient increase of KLF6 expression during trophoblast syncytialization

Pregnancy-specific Glycoprotein 1 Induces Endothelial Tubulogenesis through Interaction with Cell Surface Proteoglycans*

Pregnancy-specific β1 glycoproteins (PSGs) are the most abundant fetal proteins in the maternal bloodstream in late pregnancy. They are secreted by the syncytiotrophoblast and are detected around day 14 postfertilization. There are 11 human PSG genes, which encode a family of proteins exhibiting significant conservation at the amino acid level. We and others have proposed that PSGs have an immune modulatory function. In addition, we recently postulated that they are proangiogenic due to their ability to induce the secretion of VEGF-A and the formation of tubes by endothelial cells. The cellular receptor(s) for human PSGs remain unknown. Therefore, we conducted these studies to identify the receptor for PSG1, the highest expressed member of the family. We show that removal of cell surface glycosaminoglycans (GAGs) by enzymatic or chemical treatment of cells or competition with heparin completely inhibited binding of PSG1. In addition, PSG1 did not bind to cells lacking heparan or chondroitin sulfate on their surface, and binding was restored upon transfection with all four syndecans and glypican-1. Importantly, the presence of GAGs on the surface of endothelial cells was required for the ability of PSG1 to induce tube formation. This finding indicates that the PSG1-GAG interaction mediates at least some of the PSG1 proposed functions