PhD Scientific Days 2026

Poster Session 2.C - Molecular Medicine

Thioredoxin System Absent in Endoplasmic Reticulum Lumen: Implications for Redox Homeostasis

Name of the presenter

Veszelyi, Krisztina Nóra

Institute/workplace of the presenter

Institute of Clinical Pathophysiology

Authors

Dr. Krisztina Veszelyi¹
1: Institute of Clinical Pathophysiology

Text of the abstract

Introduction: The endoplasmic reticulum (ER) is the primary site of protein thiol oxidation and is therefore generally considered to be an oxidative environment. However, reducing processes requiring NAD(P)H are also present. In most cellular compartments, the thiol/disulfide and reduced/oxidized pyridine nucleotide systems are enzymatically linked to form complex redox networks. However, the parallel presence of oxidized thiol-disulfides and reduced pyridine nucleotides suggests that the ER lumen may lack elements that link these two systems together.
Aims: We wanted to confirm the luminal deficiency of one of these coupling systems, the thioredoxin (Trx) / thioredoxin reductase (TrxR) proteins, and to investigate the effects of their artificial expression.
Methods: We performed an in silico analysis to predict the localization of each Trx/TrxR isoform. We examined the protein expression of each isoform in subcellular fractions by Western blot analysis. Their intracellular distribution was confirmed by immunofluorescence microscopy. TrxR activity was measured using a colorimetric assay in various organelles. The cell viability of transiently transfected cells expressing Trx1 and TrxR1 in the ER lumen was measured, and the induction of apoptosis was examined by Western blot.
Results: In silico predictions showed a uniformly very low probability of luminal localization for each isoform (0–5%). Analysis of rat liver subcellular fractions revealed that none of the Trx/TrxR isoforms are expressed in the ER. Immunofluorescence analysis confirmed that Trx and TrxR isoforms did not colocalize with the ER-marker Grp94. The specific activity of TrxR in the ER was nearly zero (0,02 U/mg ± 0,01), while we measured higher activities in the cytoplasm (1,26 U/mg ± 0,11) and mitochondria (1,57 U/mg ± 0,19). Transient transfection of HeLa cells with ER-targeted Trx1 and TrxR1 resulted in a rapid decrease in cell viability and induction of apoptosis.
Conclusions: Our results suggest that none of the components of the Trx/TrxR system are expressed in the ER lumen. The absence of this electron transfer chain may explain the uncoupling of the luminal redox systems, allowing the parallel occurrence of a reduced pyridine nucleotide pool and oxidized proteins.
Funding: Semmelweis University Predoctoral Scholarship