Krüppel-like factor 6 transcription factor is required for human trophoblast fusion

A. RACCA; RIDANO, ME; CAMOLOTTO S; LIENDO M.; GENTI DE RAIMONDI S; PANZETTA DE DUTARI GM

Simposio; VI Latin American Symposium on Maternal-Fetal Interaction and Placenta (VI SLIMP)- V Latin American Symposium on Reproductive Immunology (V LASRI); 2015

SLIMP

The fusion of trophoblast cells into a highly specialized multinucleated syncytiotrophoblast layer is a key process of human placenta development. Defects in this process are associated to pathologies like preeclampsia and intrauterine growth restriction. Krüppel-Like Factor 6 (KLF6) is a member of the Sp/KLF family mostly implicated in human carcinogenesis, cell cycle regulation, and in some differentiation processes. We have previously demonstrated its expression throughout the in-vitro syncytialization process. Objective: The aim of this work was to address KLF6 contribution to syncytiotrophoblast formation by loss- and gain-of function studies. Methods and Results: KLF6 knockdown, through small interfering RNA experiments, resulted in an evident hindrance in cell-cell fusion revealed by immunofluorescence microscopy in human primary villous cytotrophoblast from term placentas as well as in the human placental-derived BeWo cell line. Moreover, KLF6 silencing led to a decrease in the expression of the fusogenic protein Syncytin-1 and the cell cycle regulator p21Cip1/Waf1, measured by quantitative RT-PCR and western blot assays. On the contrary, transcript levels of genes that encode for proteins involved in STB formation like Syncytin-1, Syncytin-2, Connexin-43, and Zonula Occludens-1 increased when KLF6 was overexpressed in the non-fusing placental-derived JEG-3 cells. Furthermore, KLF6 overexpression increased Syncytin-2 and Connexin-43 mRNAs even in villous cytotrophoblasts undergoing in-vitro syncytialization. Interestingly, the expression of two trophoblast biochemical differentiation markers, βhCG and PSG3, were not reduced after KLF6 silencing in the differentiating trophoblast cells.Conclusions: Present results support the notion that KLF6 is a relevant participant in cytotrophoblast fusion. In addition, they provide further evidence to sustain that trophoblast fusion is not a pre-requisite for biochemical differentiation.Supported by CONICET, FONCyT and SECyT