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

RACCA, ANA C.; RIDANO, MAGALÍ EVELIN; CAMOLOTTO, SOLEDAD; GENTI-RAIMONDI, SUSANA; PANZETTA-DUTARI, GRACIELA

Mar del Plata

Congreso; SLIMP-LASRI 2015; 2015

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 with 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 gain- and loss-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. In contrast, 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, was not reduced after KLF6 silencing in the differentiating trophoblast cells.Conclusions: The 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.