Mitochondrial influence in the gene regulation involved with
aging in Saccharomyces cerevisiae submitted high hydrostatic pressure.

Name: Brigida de Almeida Amorim Spagnol
Type: PhD thesis
Publication date: 07/02/2020
Advisor:

Namesort descending Role
Patricia Machado Bueno Fernandes (M/D) Advisor *

Examining board:

Namesort descending Role
Alexandre Martins Costa Santos Internal Examiner *
Antonio Alberto Ribeiro Fernandes Internal Examiner *
Monica Montero Lomeli External Examiner *
Nadja Cristhina de Souza Pinto External Examiner *
Patricia Machado Bueno Fernandes (M/D) Advisor *

Summary: Saccharomyces cerevisiae is a well-accepted eukaryotic model organism for studies on stress response and aging, WHEREas preserves several biochemical, molecular and organelle functioning mechanisms. Mitochondria is an organelle involved with stress response and, because it is main generator of reactive oxygen species (ROS) is strongly implicated with aging. Dysfunctionalities of this organelle has been associated with neurodegenerative, psychiatric, and cancer diseases. Mature and young S. cerevisiae cells respond differently to high hydrostatic pressure (HHP) stress. Cell maturity favors genes regulation involved with aging, increasing by almost three times the HHP stress resistance of these cells compared to young cells. Aging regulatory genes such as TOR1, RAS2 and SCH9 have been implicated with mitochondrial dysfunctions. However, it is possible that mitochondria influence the aging-related genes regulation. Thus, this work sought to modulate the TOR1, RAS2 and SCH9 regulation by controlling the increase in mitochondrial membrane potential (ΔΨm) of S. cerevisiae submitted to HHP. To achieve these goals, S. cerevisiae mother cells were differentiated from daughter cells by Calcofluor White impregnation into their cell wall, generating daughter cells without impregnation. Thus, ROS accumulation and ΔΨm in response to HHP was quantified by flow cytometry. Protocatechuic acid (PCA) was previously used for mitochondrial protection against the HHP unfavorable effects. Fluorescence microscopy images on mitotracker greenFM labeled cells were performed to observe mitochondrial dynamics. Finally, genes involved with aging and antioxidant defenses were analyzed by real-time PCR (qRTPCR) in the general population. Mother and daughter cells indicatived similarities in ΔΨm upon the applied treatments. Mild HHP stress indicated ΔΨm increased and 15 min recovery at ambient pressure are required for ΔΨm stabilization before severe HHP treatment. Mitochondrial protection indicated reducing of ΔΨm and this seems to have improved the tolerance to severe treatment. Mature cells accumulated high levels of ROS after HHP. However, mitochondrial protection together with the PCA antioxidant action reduced the ROS accumulation in these cells, compensating the inhibition of antioxidant genes regulation. Almost all cells with mitochondrial protection resisted to severe HHP stress. There was a 71% to 92% increase in HHP stress tolerance with previous mitochondrial protection. The genes TOR1, RAS2 and SCH9 were repressed, favoring the stress response, increasing the resistance. In addition, reducing ROS accumulation has retarded aging.

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