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DRUGPATH - a novel bioinformatic approach identifies DNA-damage pathway as a regulator of size maintenance in human ESCs and iPSCs.

ABSTRACT: Genetic and biochemical screening approaches often fail to identify functionally relevant pathway networks because many signaling proteins contribute to multiple gene ontology pathways. We developed a DRUGPATH-approach to predict pathway-interactomes from high-content drug screen data. DRUGPATH is based upon combining z-scores of effective inhibitors with their corresponding and validated targets. We test DRUGPATH by comparing homeostatic pathways in human embryonic stem cells (hESCs), human induced pluripotent stem cells (hiPSCs) and human amniotic fluid stem cells (hAFSCs). We show that hAFSCs utilize distinct interactomes compared to hESCs/hiPSCs and that pathways orchestrating cell cycle and apoptosis are strongly interconnected, while pathways regulating survival and size are not. Interestingly, hESCs/hiPSCs regulate their size by growing exact additional sizes during each cell cycle. Chemical and genetic perturbation studies show that this "adder-model" is dependent on the DNA-damage pathway. In the future, the DRUGPATH-approach may help to predict novel pathway interactomes from high-content drug screens.

SUBMITTER: Kovacic B

PROVIDER: S-EPMC6374489 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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DRUGPATH - a novel bioinformatic approach identifies DNA-damage pathway as a regulator of size maintenance in human ESCs and iPSCs.

Kovacic Boris B Rosner Margit M Schlangen Karin K Kramer Nina N Hengstschläger Markus M

Scientific reports 20190213 1

Genetic and biochemical screening approaches often fail to identify functionally relevant pathway networks because many signaling proteins contribute to multiple gene ontology pathways. We developed a DRUGPATH-approach to predict pathway-interactomes from high-content drug screen data. DRUGPATH is based upon combining z-scores of effective inhibitors with their corresponding and validated targets. We test DRUGPATH by comparing homeostatic pathways in human embryonic stem cells (hESCs), human ind ...[more]

PMID: 30760778

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Project description:Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have distinct origins: ESCs are derived from pre-implanted embryos while iPSCs are reprogrammed somatic cells. Both have their own characteristics and lineage specificity, and both are valuable tools for studying human neurological development and disease. Thus far, few studies have analyzed how differences between stem cell types influence mitochondrial function and mitochondrial DNA (mtDNA) homeostasis during differentiation into neural and glial lineages. In this study, we compared mitochondrial function and mtDNA replication in human ESCs and iPSCs at three different stages - pluripotent, neural progenitor and astrocyte. We found that while ESCs and iPSCs have a similar mitochondrial signature, neural and astrocyte derivations manifested differences. At the neural stem cell (NSC) stage, iPSC-NSCs displayed decreased ATP production and a reduction in mitochondrial respiratory chain (MRC) complex IV expression compared to ESC-NSCs. IPSC-astrocytes showed increased mitochondrial activity including elevated ATP production, MRC complex IV expression, mtDNA copy number and mitochondrial biogenesis relative to those derived from ESCs. These findings show that while ESCs and iPSCs are similar at the pluripotent stage, differences in mitochondrial function may develop during differentiation and must be taken into account when extrapolating results from different cell types.Abbreviation: BSA: Bovine serum albumin; DCFDA: 2',7'-dichlorodihydrofluorescein diacetate; DCX: Doublecortin; EAAT-1: Excitatory amino acid transporter 1; ESCs: Embryonic stem cells; GFAP: Glial fibrillary acidic protein; GS: Glutamine synthetase; iPSCs: Induced pluripotent stem cells; LC3B: Microtubule-associated protein 1 light chain 3β; LC-MS: Liquid chromatography-mass spectrometry; mito-ROS: Mitochondrial ROS; MMP: Mitochondrial membrane potential; MRC: Mitochondrial respiratory chain; mtDNA: Mitochondrial DNA; MTDR: MitoTracker Deep Red; MTG: MitoTracker Green; NSCs: Neural stem cells; PDL: Poly-D-lysine; PFA: Paraformaldehyde; PGC-1α: PPAR-γ coactivator-1 alpha; PPAR-γ: Peroxisome proliferator-activated receptor-gamma; p-SIRT1: Phosphorylated sirtuin 1; p-ULK1: Phosphorylated unc-51 like autophagy activating kinase 1; qPCR: Quantitative PCR; RT: Room temperature; RT-qPCR: Quantitative reverse transcription PCR; SEM: Standard error of the mean; TFAM: Mitochondrial transcription factor A; TMRE: Tetramethylrhodamine ethyl ester; TOMM20: Translocase of outer mitochondrial membrane 20.

Dataset Information

DRUGPATH - a novel bioinformatic approach identifies DNA-damage pathway as a regulator of size maintenance in human ESCs and iPSCs.

Publications

DRUGPATH - a novel bioinformatic approach identifies DNA-damage pathway as a regulator of size maintenance in human ESCs and iPSCs.

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