TFEB controls expression of human syncytins during cell-cell fusion [ChIP-Seq]

During human development, a subset of differentiating fetal cells form a temporary organ, the placenta, which invades the uterine wall to support nutrient, oxygen, and waste exchange between the mother and fetus until birth. Most of the human placenta is formed by a syncytial villous structure which arises via cell-cell fusion of underlying fetal trophoblast stem cells. Genetic and functional studies have characterized the membrane protein fusogens, Syncytin-1 and Syncytin-2, that are both necessary and sufficient for human trophoblast cell-cell fusion. However, identification and characterization of upstream transcriptional regulators regulating their expression has been limited. Here, using CRISPR knockout in an in vitro cellular model of syncytiotrophoblast development (BeWo cells), we find that the transcription factor TFEB, mainly known as a regulator of autophagy and lysosomal biogenesis, is required for cell-cell fusion of syncytiotrophoblasts. TFEB translocates to the nucleus, exhibits increased chromatin interactions, and directly binds the Syncytin-1 and Syncytin-2 promoters to control their expression during differentiation. While TFEB appears to play an important role in syncytiotrophoblast differentiation, ablation of TFEB largely does not affect lysosomal gene expression or lysosomal biogenesis in differentiating BeWo cells, suggesting that TFEB plays an alternative role in placental cells. Overall design: We examined the binding of TFEB to genomic loci using ChIP-sequencing during syncytiotrophoblast differentiation in BeWo cells. We compared binding of TFEB in BeWo cells stably expressing 3xFLAG-Halo-TFEB under a low expression promoter (L30) and an antibiotic cassette (IRES Puro). We compared genomic enrichment after immunoprecipatation using an anti-FLAG antibody versus IgG pulldown or Input samples before precipitation. We compared TFEB localization to specific genes after treatment with DMSO, Forskolin-induced differentiation, or after treatment with the mTOR inhibitor Torin1.