2. Academic Validation
  3. Temporal dynamics of proteome and phosphorproteome during neuronal differentiation in the reference KOLF2.1J iPSC line

Temporal dynamics of proteome and phosphorproteome during neuronal differentiation in the reference KOLF2.1J iPSC line

  • bioRxiv. 2025 Mar 26:2025.03.25.645331. doi: 10.1101/2025.03.25.645331.
Ying Hao 1 2,  Ziyi Li 1 3 2,  Erika Lara 1,  Daniel M Ramos 1,  Marianist Santiana 1,  Benjamin Jin 1,  Jacob Epstein 1,  Jasmin Camacho 1,  Nicole Carmiol 1,  Isabelle Kowal 1,  Paige Jarreau 1,  Cory A Weller 1 3,  Sydney Klaisner 1,  Laurel A Screven 1,  Caroline B Pantazis 1,  Mike A Nalls 1 3,  Priyanka Narayan 1 4,  Luigi Ferrucci 5,  Andrew B Singleton 1 6,  Michael E Ward 7,  Mark R Cookson 1 6,  Yue Andy Qi 1
Affiliations

Affiliations

  • 1 Center for Alzheimer's and Related Dementias (CARD), National Institute on Aging and National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD, USA.
  • 2 These authors contributed equally.
  • 3 DataTecnica LLC, Washington, DC, USA.
  • 4 Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA.
  • 5 Longitudinal Studies Section, Translational Gerontology Branch, National Institute on Aging, Baltimore, MD, USA.
  • 6 Laboratory of Neurogenetics, National Institute on Aging, National Institutes of Health, Bethesda, MD, USA.
  • 7 National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD, USA.
PMID: 40568166 DOI: 10.1101/2025.03.25.645331
Abstract

Induced pluripotent stem cell (iPSC)-derived neurons have emerged as a powerful model to investigate both neuronal development and neurodegenerative diseases. Although transcriptomics and imaging have been applied to characterize neuronal development signatures, comprehensive datasets of protein and post-translational modifications (PTMs) are not readily available. Here, we applied quantitative proteomics and phosphoproteomics to profile the differentiation of the KOLF2.1J iPSC line, the first reference line of the iPSC Neurodegenerative Disease Initiative (iNDI) project. We developed an automated workflow enabling high-coverage enrichment of proteins and phosphoproteins. Our results revealed molecular signatures across proteomic and phosphoproteomic landscapes during differentiation of iPSC-derived neurons. Proteomic data highlighted distinct changes in mitochondrial pathways throughout the course of differentiation, while phosphoproteomics revealed specific regulatory dynamics in GTPase signaling pathways and microtubule proteins. Additionally, phosphosite dynamics exhibited discordant trends compared to protein expression, particularly in processes related to axon functions and RNA transport. Furthermore, we mapped the kinase dynamic changes that are critical for neuronal development and maturation. We developed an interactive Web app (https://niacard.shinyapps.io/Phosphoproteome/) to visualize temporal landscape dynamics of protein and phosphosite expression. By establishing baselines of proteomic and phosphoproteomic profiles for neuronal differentiation, this dataset offers a valuable resource for future research into neuronal development and neurodegenerative diseases using this reference iPSC line.

Keywords

GTPase; KOLF2.1J; iNDI; kinase; microtubule; mitochondrial; neuronal differentiation; phosphoproteomics; proteomics.

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