Mapping prohormone processing by proteases in human enteroendocrine cells using genetically engineered organoid models

Joep Beumer, Julia Bauzá-Martinez, Tim S. Veth, Veerle Geurts, Charelle Boot, Hannah Gilliam-Vigh, Steen S. Poulsen, Filip K. Knop, Wei Wu, Hans Clevers

1 Citation (Scopus)

Abstract

Enteroendocrine cells (EECs) secrete hormones in response to ingested nutrients to control physiological processes such as appetite and insulin release. EEC hormones are synthesized as large proproteins that undergo proteolytic processing to generate bioactive peptides. Mutations in EEC-enriched proteases are associated with endocrinopathies. Due to the relative rarity of EECs and a paucity of in vitro models, intestinal prohormone processing remains challenging to assess. Here, human gut organoids in which EECs can efficiently be induced are subjected to CRISPR-Cas9-mediated modification of EEC-expressed endopeptidase and exopeptidase genes. We employ mass spectrometry-based analyses to monitor peptide processing and identify glucagon production in intestinal EECs, stimulated upon bone morphogenic protein (BMP) signaling. We map the substrates and products of major EECs endo- and exopeptidases. Our studies provide a comprehensive description of peptide hormones produced by human EECs and define the roles of specific proteases in their generation.

Original languageEnglish
Article numbere2212057119
JournalProceedings of the National Academy of Sciences of the United States of America
Volume119
Issue number46
Pages (from-to)e2212057119
ISSN0027-8424
DOIs
Publication statusPublished - 16 Nov 2022

Keywords

  • CRISPR-Cas9
  • enteroendocrine cells
  • intestinal organoids
  • peptidomics
  • prohormone processing

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