Author:

Keywords:

Science & Technology, Life Sciences & Biomedicine, Cell & Tissue Engineering, Biotechnology & Applied Microbiology, Oncology, Cell Biology, Hematology, cell state transitions, embryonic stem cells, esiRNA, genetic interaction, neural differentiation, signaling pathway, TGF beta, BMP, SELF-RENEWAL, PLURIPOTENCY FACTORS, BMP4 SYNEXPRESSION, HUMAN ES, DIFFERENTIATION, EXPRESSION, FATE, GENES, NANOG, ZEB2, TGFβ/BMP, Bone Morphogenetic Proteins, Cell Differentiation, Embryonic Stem Cells, Humans, Transforming Growth Factor beta, 06 Biological Sciences, 10 Technology, 11 Medical and Health Sciences, Immunology, 31 Biological sciences, 32 Biomedical and clinical sciences

Abstract:

Cooperative actions of extrinsic signals and cell-intrinsic transcription factors alter gene regulatory networks enabling cells to respond appropriately to environmental cues. Signaling by transforming growth factor type β (TGFβ) family ligands (eg, bone morphogenetic proteins [BMPs] and Activin/Nodal) exerts cell-type specific and context-dependent transcriptional changes, thereby steering cellular transitions throughout embryogenesis. Little is known about coordinated regulation and transcriptional interplay of the TGFβ system. To understand intrafamily transcriptional regulation as part of this system's actions during development, we selected 95 of its components and investigated their mRNA-expression dynamics, gene-gene interactions, and single-cell expression heterogeneity in mouse embryonic stem cells transiting to neural progenitors. Interrogation at 24 hour intervals identified four types of temporal gene transcription profiles that capture all stages, that is, pluripotency, epiblast formation, and neural commitment. Then, between each stage we performed esiRNA-based perturbation of each individual component and documented the effect on steady-state mRNA levels of the remaining 94 components. This exposed an intricate system of multilevel regulation whereby the majority of gene-gene interactions display a marked cell-stage specific behavior. Furthermore, single-cell RNA-profiling at individual stages demonstrated the presence of detailed co-expression modules and subpopulations showing stable co-expression modules such as that of the core pluripotency genes at all stages. Our combinatorial experimental approach demonstrates how intrinsically complex transcriptional regulation within a given pathway is during cell fate/state transitions.