Jud, A. | bioRxiv Science

bioRxiv2024

Characterization of the Glutathione Redox State in the Golgi Apparatus

Redox homeostasis is crucial for cell function, and, in eukaryotic cells, studying it in a compartmentalized way is essential due to the redox variations between different organelles. The redox state of organelles is largely determined by the redox potential of glutathione, EGSH, and the concentration of its reduced and oxidized species, [GS]. The Golgi apparatus is an essential component of the secretory pathway, yet little is known about the concentration or redox state of GSH in this organelle. Here, we characterized the redox state of GSH in the Golgi apparatus using a combination of microscopy and proteomics methods. Our results prove that the Golgi apparatus is a highly oxidizing organelle with a strikingly low GSH concentration (EGSH = - 157 mV, 1-5 mM). These results fill an important gap in our knowledge of redox homeostasis in subcellular organelles. Moreover, the new Golgi-targeted GSH sensors allow us to observe dynamic changes in the GSH redox state in the organelle and pave the way for robust characterization of the Golgi redox state under various physiological and pathological conditions.

cell biology↗

bioRxiv2021

Proteome biology of primary colorectal carcinoma and corresponding liver metastases

Colorectal adenocarcinomas (CRC) are one of the most commonly diagnosed tumors worldwide. Colorectal adenocarcinomas primarily metastasize into the liver and (less often) into the peritoneum. Patients suffering from CRC-liver metastasis (CRC-LM) typically present with a dismal overall survival compared to non-metastasized CRC patients. The metastasis process and metastasis-promoting factors in patients with CRC are under intensive debate. However, CRC studies investigating the proteome biology are lacking. Formalin-fixed paraffin-embedded (FFPE) tissue specimens provide a valuable resource for comprehensive proteomic studies of a broad variety of clinical malignancies. The presented pilot study compares the proteome of primary CRC and patient-matched CRC-LM. The applied protocol allows a reproducible and straightforward identification and quantification of over 2,600 proteins within the dissected tumorous tissue. Subsequent unsupervised clustering reveals distinct proteome biologies of the primary CRC and the corresponding CRC-LM. Statistical analysis yields multiple differentially abundant proteins in either primary CRC or their corresponding liver metastases. A more detailed analysis of dysregulated biological processes suggests an active immune response in the liver metastases, including several proteins of the complement system. Proteins with structural roles, e.g. cytoskeleton organization or cell junction assembly appear to be less prominent in liver metastases as compared to primary CRC. Immunohistochemistry corroborates proteomic high expression levels of metabolic proteins in CRC-LM. We further assessed how the in vitro inhibition of two in CRC-LM enriched metabolic proteins affected cell proliferation and chemosensitivity. The presented proteomic investigation in a small clinical cohort promotes a more comprehensive understanding of the distinct proteome biology of primary CRC and their corresponding liver metastases.

cancer biology↗