EP300 facilitates human trophoblast stem cell differentiation.

Early placenta development involves cytotrophoblast differentiation into extravillous trophoblast (EVT) and syncytiotrophoblast (STB). Defective trophoblast development and function may result in severe pregnancy complications, including fetal growth restriction and pre-eclampsia. The incidence of these complications is increased in pregnancies of fetuses affected by Rubinstein-Taybi syndrome, a developmental disorder predominantly caused by heterozygous mutations in CREB-binding protein (CREBBP) or E1A-binding protein p300 (EP300). Although the acetyltransferases CREBBP and EP300 are paralogs with many overlapping functions, the increased incidence of pregnancy complications is specific for EP300 mutations. We hypothesized that these complications have their origin in early placentation and that EP300 is involved in that process. Therefore, we investigated the role of EP300 and CREBBP in trophoblast differentiation, using human trophoblast stem cells (TSCs) and trophoblast organoids. We found that pharmacological CREBBP/EP300 inhibition blocks differentiation of TSCs into both EVT and STB lineages, and results in an expansion of TSC-like cells under differentiation-inducing conditions. Specific targeting by RNA interference or CRISPR/Cas9-mediated mutagenesis demonstrated that knockdown of EP300 but not CREBBP, inhibits trophoblast differentiation, consistent with the complications seen in Rubinstein-Taybi syndrome pregnancies. By transcriptome sequencing, we identified transforming growth factor alpha (TGFA, encoding TGF-α) as being strongly upregulated upon EP300 knockdown. Moreover, supplementing differentiation medium with TGF-α, which is a ligand for the epidermal growth factor receptor (EGFR), likewise affected trophoblast differentiation and resulted in increased TSC-like cell proliferation. These findings suggest that EP300 facilitates trophoblast differentiation by interfering with at least EGFR signaling, pointing towards a crucial role for EP300 in early human placentation.

TBX2, a Novel Regulator of Labour.

Background and Objectives: Therapeutic interventions targeting molecular factors involved in the transition from uterine quiescence to overt labour are not substantially reducing the rate of spontaneous preterm labour. The identification of novel rational therapeutic targets are essential to prevent the most common cause of neonatal mortality. Based on our previous work showing that Tbx2 (T-Box transcription factor 2) is a putative upstream regulator preceding progesterone withdrawal in mouse myometrium, we now investigate the role of TBX2 in human myometrium. Materials and Methods: RNA microarray analysis of (A) preterm human myometrium samples and (B) myometrial cells overexpressing TBX2 in vitro, combined with subsequent analysis of the two publicly available datasets of (C) Chan et al. and (D) Sharp et al. The effect of TBX2 overexpression on cytokines/chemokines secreted to the myometrium cell culture medium were determined by Luminex assay. Results: Analysis shows that overexpression of TBX2 in myometrial cells results in downregulation of TNFα- and interferon signalling. This downregulation is consistent with the decreased expression of cytokines and chemokines of which a subset has been previously associated with the inflammatory pathways relevant for human labour. In contrast, CXCL5 (C-X-C motif chemokine ligand 5), CCL21 and IL-6 (Interleukin 6), previously reported in relation to parturition, do not seem to be under TBX2 control. The combined bioinformatical analysis of the four mRNA datasets identifies a subset of upstream regulators common to both preterm and term labour under control of TBX2. Surprisingly, TBX2 mRNA levels are increased in preterm contractile myometrium. Conclusions: We identified a subset of upstream regulators common to both preterm and term labour that are activated in labour and repressed by TBX2. The increased TBX2 mRNA expression in myometrium collected during a preterm caesarean section while in spontaneous preterm labour compared to tissue harvested during iatrogenic preterm delivery does not fit the bioinformatical model. We can only explain this by speculating that the in vivo activity of TBX2 in human myometrium depends not only on the TBX2 expression levels but also on levels of the accessory proteins necessary for TBX2 activity.

Genes differentially expressed in thyroid carcinoma identified by comparison of SAGE expression profiles.

To identify transcripts that distinguish malignant from benign thyroid disease serial analysis of gene expression (SAGE) profiles of papillary thyroid carcinoma and of normal thyroid are compared. Of the 21,000 tags analyzed, 204 tags are differentially expressed with statistical significance in the tumor. Thyroid tumor specificity of these transcripts is determined in silico using the tissue preferential expression (TPE) algorithm. TPE values demonstrate that 42 tags of the 204 are thyroid tumor specific. BC013035, a cDNA encoding a novel protein, is up-regulated from 0 to 24 tags in the thyroid tumor SAGE library. In a tissue panel of 30 thyroid tumors and 12 controls, it has an expression pattern similar to thyroid peroxidase, indicating possible involvement of BC013035 in thyroid differentiation. A tag coding for extracellular matrix protein 1 (ECM1) is absent in the normal thyroid SAGE library and present 55 times in the tumor. ECM1, a protein recently associated with angiogenesis and expressed in metastatic breast carcinoma, is up-regulated in 50% of all thyroid carcinoma and absent in normal controls and follicular adenoma. In conclusion, SAGE analysis and subsequent determination of TPE values facilitates the rapid distinction of genes specifically expressed in cancer tissues.