The Role of Hydrogen Peroxide on Nuclear Actin Dynamics in Hypoxia

by Carla Marie Siemssen
Doctoral thesis
Date of Examination:2024-10-22
Date of issue:2024-09-13
Advisor:PD Dr. Anke Zieseniß
Referee:PD Dr. Anke Zieseniß
Referee:PD Dr. Elisabeth Heßmann
Persistent Address: http://dx.doi.org/10.53846/goediss-10738

 

 

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Abstract

English

With an increasing interest in the dynamics of the highly conserved protein actin and the signaling function of reactive oxygen species, this thesis aimed to discover if changes in reactive oxygen species production, specifically hydrogen peroxide, are the cause of the fast depletion of monomeric globular actin (G-actin) observed in various cell lines after the onset of hypoxia. To study the effect of endogenous hydrogen peroxide production on the nuclear G-actin content of the cell, Hep3B cells were transduced with the HyPer-DAO fusion protein, a powerful tool for externally controlled hydrogen peroxide production and simultaneous monitoring. The newly established Hep3B-HyPer-DAO-NES and -NLS cell lines showed precise localization of the construct in either the nucleus (NLS) or the cytoplasm (NES) and appeared to function without limitations in normoxia and hypoxia during live cell HyPer measurements. Results employing the established HyPer-DAO cell clones demonstrated that the endogenous production of hydrogen peroxide in neither the nucleus nor the cytoplasm induced nuclear G-actin depletion in normoxia and furthermore, did not prevent the deceleration of nuclear G-actin levels in hypoxia. Hence, until now, hydrogen peroxide does not seem to profoundly influence the fast-changing dynamics of nuclear actin monomers after the onset of hypoxia.
Keywords: actin; hypoxia; hydrogen peroxide; HyPer-DAO; actin dynamics