ChIP-Seq and RNA-Seq analyses identify components of the Wnt and Fgf signaling pathways as Prep1 target genes in mouse embryonic stem cells

Audrey Laurent; Manuela Calabrese; Hans-Jörg Warnatz; Marie-Laure Yaspo; Vsevolod Tkachuk; Miguel Torres; Francesco Blasi; Dmitry Penkov

doi:10.1371/journal.pone.0122518

ChIP-Seq and RNA-Seq analyses identify components of the Wnt and Fgf signaling pathways as Prep1 target genes in mouse embryonic stem cells

PLoS One. 2015 Apr 13;10(4):e0122518. doi: 10.1371/journal.pone.0122518. eCollection 2015.

Authors

Audrey Laurent¹, Manuela Calabrese¹, Hans-Jörg Warnatz², Marie-Laure Yaspo², Vsevolod Tkachuk³, Miguel Torres⁴, Francesco Blasi¹, Dmitry Penkov⁵

Affiliations

¹ IFOM (FIRC Institute of Molecular Oncology), IFOM-IEO-Campus, Milan, Italy.
² Department of Vertebrate Genomics, Max Planck Institute for Molecular Genetics, Berlin, Germany.
³ Faculty of Basic Medicine, Lomonosov Moscow State University, Moscow, Russia.
⁴ Department of Cardiovascular Development and Repair, Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain.
⁵ IFOM (FIRC Institute of Molecular Oncology), IFOM-IEO-Campus, Milan, Italy; Department of Experimental Cardiology, Russian Cardiology Research and Production Complex, Moscow, Russia.

Abstract

The Prep1 (Pknox1) homeodomain transcription factor is essential at multiple stages of embryo development. In the E11.5 embryo trunk, we previously estimated that Prep1 binds about 3,300 genomic sites at a highly specific decameric consensus sequence, mainly governing basal cellular functions. We now show that in embryonic stem (ES) cells Prep1 binding pattern only partly overlaps that of the embryo trunk, with about 2,000 novel sites. Moreover, in ES cells Prep1 still binds mostly to promoters, as in total embryo trunk but, among the peaks bound exclusively in ES cells, the percentage of enhancers was three-fold higher. RNA-seq identifies about 1800 genes down-regulated in Prep1-/- ES cells which belong to gene ontology categories not enriched in the E11.5 Prep1i/i differentiated embryo, including in particular essential components of the Wnt and Fgf pathways. These data agree with aberrant Wnt and Fgf expression levels in the Prep1-/- ES cells with a deficient embryoid bodies (EBs) formation and differentiation. Re-establishment of the Prep1 level rescues the phenotype.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Binding Sites
Cell Differentiation / genetics*
DNA-Binding Proteins / biosynthesis*
DNA-Binding Proteins / genetics
Embryo, Mammalian
Embryonic Development / genetics
Fibroblast Growth Factors / biosynthesis
Gene Expression Regulation, Developmental
Genome
Homeodomain Proteins / biosynthesis*
Homeodomain Proteins / genetics
Mice
Mouse Embryonic Stem Cells*
Wnt Signaling Pathway / genetics

Substances

DNA-Binding Proteins
Homeodomain Proteins
Pknox1 protein, mouse
Fibroblast Growth Factors

Associated data

GEO/GSE63282

Grants and funding

Without the support of COST BM0805 action and the ensuing inter-laboratory exchanges, this work would have not been possible. AL was supported by a fellowship of FUV (Fondazione Umberto Veronesi). FB was supported by AIRC (Italian Association for Cancer Research, grant n. BM0805), Cariplo Foundation and Italian Ministry of Health. JIW and MLY were supported by the Max Planck Society (Munich, Germany) and the European Commission under its sixth Framework Programme (FP6, grant AnEUploidy [LSHG-CT-2006-037627]). Work in MT laboratory was supported by grant BFU2012-31086 from the Spanish Ministerio de Economía y Competitividad. DP was funded from the Russian Foundation for Basic Research (12-04-01659-а); VT was supported by Russian Research Foundation Grant (project 14-24-00086). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.