Procathepsin V is secreted in a TSH regulated manner from human thyroid epithelial cells and is accessible to an activity-based probe

Alaa Al-Hashimi, Vaishnavi Venugopalan, Maren Rehders, Naphannop Sereesongsaeng, Zeynep Hein, Sebastian Springer, Ekkehard Weber, Dagmar Führer, Matthew S. Bogyo, Christopher J. Scott, Roberta E. Burden, Klaudia Brix*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

5 Citations (Scopus)
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Abstract

The significance of cysteine cathepsins for the liberation of thyroid hormones from the precursor thyroglobulin was previously shown by in vivo and in vitro studies. Cathepsin L is most important for thyroglobulin processing in mice. The present study aims at specifying the possible contribution of its closest relative, cysteine cathepsin L2/V, to thyroid function. Immunofluorescence analysis on normal human thyroid tissue revealed its predominant localization at the apical plasma membrane of thyrocytes and within the follicle lumen, indicating the secretion of cathepsin V and extracellular tasks rather than its acting within endo-lysosomes. To explore the trafficking pathways of cathepsin V in more detail, a chimeric protein consisting of human cathepsin V tagged with green fluorescent protein (GFP) was stably expressed in the Nthyori 3-1 thyroid epithelial cell line. Colocalization studies with compartment-specific markers and analyses of post-translational modifications revealed that the chimeric protein was sorted into the lumen of the endoplasmic reticulum and subsequently transported to the Golgi apparatus, while being N-glycosylated. Immunoblotting showed that the chimeric protein reached endo-lysosomes and it became secreted from the transduced cells. Astonishingly, thyroid stimulating hormone (TSH)-induced secretion of GFP-tagged cathepsin V occurred as the proform, suggesting that TSH upregulates its transport to the plasma membrane before it reaches endo-lysosomes for maturation. The proform of cathepsin V was found to be reactive with the activity-based probe DCG-04, suggesting that it possesses catalytic activity. We propose that TSH-stimulated secretion of procathepsin V is the default pathway in the thyroid to enable its contribution to thyroglobulin processing by extracellular means.

Original languageEnglish
Article number9140
Number of pages23
JournalInternational Journal of Molecular Sciences
Volume21
Issue number23
Early online date30 Nov 2020
DOIs
Publication statusPublished - 01 Dec 2020

Bibliographical note

Funding Information:
Funding: This research was funded by Deutscher Akademischer Austauschdienst (DAAD), Research Grants – Doctoral Programmes in Germany, grant number 91534725 to A.A.-H. and by Jacobs University Bremen, Germany, project number 6113/90140 to K.B. This research was further supported by Deutsche Forschungsgemeinschaft (DFG), Bonn, Germany, in the framework of the FOR 367, grant number BR 1308/6-1 and 6-2 to K.B.

Publisher Copyright:
© 2020 by the authors. Licensee MDPI, Basel, Switzerland.

Keywords

  • Cysteine cathepsins
  • Green fluorescent protein tagging
  • Protein trafficking
  • Secretion
  • Thyroid epithelial cells
  • Thyroid stimulating hormone

ASJC Scopus subject areas

  • Catalysis
  • Molecular Biology
  • Spectroscopy
  • Computer Science Applications
  • Physical and Theoretical Chemistry
  • Organic Chemistry
  • Inorganic Chemistry

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