Defective insulin receptor signaling in hPSCs skews pluripotency and negatively perturbs neural differentiation
| Autor: | Wei-Jun Qian, Chang Siang Lim, Manoj K. Gupta, Hwee Hui Lau, John L. Rinn, Marina A. Gritsenko, Larry Sai Weng Loo, William Mallard, Nicholas Jackson, Linh Nguyen, Richard D. Smith, Rohit N. Kulkarni, Adrian Kee Keong Teo |
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| Rok vydání: | 2021 |
| Předmět: |
Pluripotent Stem Cells
Proteomics 0301 basic medicine KOSR IGF insulin-like growth factor Human Embryonic Stem Cells hESC human embryonic stem cell Biology Cell fate determination Biochemistry neural lineage Cell Line 03 medical and health sciences SOX1 Downregulation and upregulation stem cells Humans human Phosphorylation Induced pluripotent stem cell Molecular Biology cell fate specification Cells Cultured Mitogen-Activated Protein Kinase 1 Neurons Mitogen-Activated Protein Kinase 3 ERK1/2 KOSR KnockOut Serum Replacement 030102 biochemistry & molecular biology Neuroectoderm AKT Cell Differentiation differentiation Cell Biology pluripotency Embryonic stem cell Receptor Insulin ECM extracellular matrix Cell biology hPSC human pluripotent stem cell 030104 developmental biology IR insulin receptor insulin receptors Stem cell signaling Octamer Transcription Factor-3 Research Article Signal Transduction |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 0021-9258 |
| Popis: | Human embryonic stem cells are a type of pluripotent stem cells (hPSCs) that are used to investigate their differentiation into diverse mature cell types for molecular studies. The mechanisms underlying insulin receptor (IR)-mediated signaling in the maintenance of human pluripotent stem cell (hPSC) identity and cell fate specification are not fully understood. Here, we used two independent shRNAs to stably knock down IRs in two hPSC lines that represent pluripotent stem cells and explored the consequences on expression of key proteins in pathways linked to proliferation and differentiation. We consistently observed lowered pAKT in contrast to increased pERK1/2 and a concordant elevation in pluripotency gene expression. ERK2 chromatin immunoprecipitation, luciferase assays, and ERK1/2 inhibitors established direct causality between ERK1/2 and OCT4 expression. Of importance, RNA sequencing analyses indicated a dysregulation of genes involved in cell differentiation and organismal development. Mass spectrometry–based proteomic analyses further confirmed a global downregulation of extracellular matrix proteins. Subsequent differentiation toward the neural lineage reflected alterations in SOX1+PAX6+ neuroectoderm and FOXG1+ cortical neuron marker expression and protein localization. Collectively, our data underscore the role of IR-mediated signaling in maintaining pluripotency, the extracellular matrix necessary for the stem cell niche, and regulating cell fate specification including the neural lineage. |
| Databáze: | OpenAIRE |
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