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1.
Int J Mol Sci ; 24(8)2023 Apr 21.
Article in English | MEDLINE | ID: mdl-37108830

ABSTRACT

One of the main inducers of autophagy-dependent self-cannibalism, called ULK1, is tightly regulated by the two sensor molecules of nutrient conditions and energy status, known as mTOR and AMPK kinases, respectively. Recently, we developed a freely available mathematical model to explore the oscillatory characteristic of the AMPK-mTOR-ULK1 regulatory triangle. Here, we introduce a systems biology analysis to explain in detail the dynamical features of the essential negative and double-negative feedback loops and also the periodic repeat of autophagy induction upon cellular stress. We propose an additional regulatory molecule in the autophagy control network that delays some of AMPK's effect on the system, making the model output more consistent with experimental results. Furthermore, a network analysis on AutophagyNet was carried out to identify which proteins could be the proposed regulatory components in the system. These regulatory proteins should satisfy the following rules: (1) they are induced by AMPK; (2) they promote ULK1; (3) they down-regulate mTOR upon cellular stress. We have found 16 such regulatory components that have been experimentally proven to satisfy at least two of the given rules. Identifying such critical regulators of autophagy induction could support anti-cancer- and ageing-related therapeutic efforts.


Subject(s)
AMP-Activated Protein Kinases , Intracellular Signaling Peptides and Proteins , AMP-Activated Protein Kinases/metabolism , Intracellular Signaling Peptides and Proteins/pharmacology , Autophagy-Related Protein-1 Homolog/genetics , Autophagy-Related Protein-1 Homolog/metabolism , Systems Biology , TOR Serine-Threonine Kinases/metabolism , Autophagy
2.
FEBS Open Bio ; 12(11): 2065-2082, 2022 11.
Article in English | MEDLINE | ID: mdl-36097827

ABSTRACT

Endoplasmic reticulum (ER) stress-dependent accumulation of incorrectly folded proteins leads to activation of the unfolded protein response. The role of the unfolded protein response (UPR) is to avoid cell damage and restore the homeostatic state by autophagy; however, excessive ER stress results in apoptosis. Here we investigated the ER stress-dependent feedback loops inside one of the UPR branches by focusing on PERK-induced ATF4 and its two targets, called CHOP and GADD34. Our goal was to qualitatively describe the dynamic behavior of the system by exploring the key regulatory motifs using both molecular and theoretical biological techniques. Using the HEK293T cell line as a model system, we confirmed that the life-or-death decision is strictly regulated. We investigated the dynamic characteristics of the crucial elements of the PERK pathway at both the RNA and protein level upon tolerable and excessive levels of ER stress. Of particular note, inhibition of GADD34 or CHOP resulted in various phenotypes upon high levels of ER stress. Our computer simulations suggest the existence of two new feedback loops inside the UPR. First, GADD34 seems to have a positive effect on ATF4 activity, while CHOP inhibits it. We claim that these newly described feedback loops ensure the fine-tuning of the ATF4-dependent stress response mechanism of the cell.


Subject(s)
Activating Transcription Factor 4 , Endoplasmic Reticulum Stress , Humans , Activating Transcription Factor 4/metabolism , HEK293 Cells , Transcription Factor CHOP/genetics , Transcription Factor CHOP/metabolism , Unfolded Protein Response
3.
Cell Death Dis ; 12(4): 309, 2021 03 23.
Article in English | MEDLINE | ID: mdl-33758166

ABSTRACT

Although autophagy is a type of programmed cell death, it is also essential for cell survival upon tolerable level of various stress events. For the cell to respond adequately to an external and/or internal stimulus induced by cellular stress, autophagy must be controlled in a highly regulated manner. By using systems biology techniques, here we explore the dynamical features of autophagy induction. We propose that the switch-like characteristic of autophagy induction is achieved by a control network, containing essential feedback loops of four components, so-called autophagy inducer, autophagy controller, mTORC1 and autophagy executor, respectively. We show how an autophagy inducer is capable to turn on autophagy in a cellular stress-specific way. The autophagy controller acts as a molecular switch and not only promotes autophagy but also blocks the permanent hyperactivation of the process via downregulating the autophagy inducer. In this theoretical analysis, we explore in detail the properties of all four proposed controlling elements and their connections. Here we also prove that the kinetic features of this control network can be considered accurate in various stress processes (such as starvation, endoplasmic reticulum stress and oxidative stress), even if the exact components may be different. The robust response of the resulting control network is essential during cellular stress.


Subject(s)
Autophagy/genetics , Cell Survival/genetics , Stress, Physiological/genetics , Humans
4.
FEBS Lett ; 594(6): 1112-1123, 2020 03.
Article in English | MEDLINE | ID: mdl-31769869

ABSTRACT

Scientific results have revealed that autophagy is able to promote cell survival in response to endoplasmic reticulum (ER) stress, while drastic events result in apoptotic cell death. Here, we analyse the important crosstalk of life-and-death decisions from a systems biological perspective by studying the regulatory modules of the unfolded protein response (UPR). While a double-negative loop between autophagy and apoptosis inducers is crucial for the switch-like characteristic of the stress response mechanism, a positive feedback loop between ER stress sensors is also essential. Corresponding to experimental data, here, we show the dynamical significance of Gadd34-CHOP connections inside the PERK branch of the UPR. The multiple system-level feedback loops seem to be crucial for managing a robust life-and-death decision depending on the level and durability of cellular stress.


Subject(s)
Apoptosis/physiology , Autophagy/physiology , Endoplasmic Reticulum Stress/physiology , Models, Biological , Unfolded Protein Response/physiology , Feedback , Humans , Protein Phosphatase 1/metabolism , Transcription Factor CHOP/metabolism
5.
PLoS One ; 13(11): e0207949, 2018.
Article in English | MEDLINE | ID: mdl-30485363

ABSTRACT

Oxidative stress results in activation of several signal transduction pathways controlled by the PERK-substrate NRF2 (nuclear factor erythroid 2-related factor 2); meanwhile the ongoing cell division cycle has to be blocked. It has been recently shown that Cyclin D1 got immediately down-regulated via PERK pathway in response to oxidative stress leading to cell cycle arrest. However, the effect of NRF2 on cell cycle regulation has not been explored yet. We aimed to reveal a crosstalk between PERK-substrate NRF2 and the key elements of cell cycle regulatory network upon oxidative stress using molecular biological techniques- Although Cyclin D1 level remained constant, its activity was blocked by various stoichiometric inhibitors (such as p15, p21 and p27) even at low level of oxidative stress. The activity of these CDK inhibitors completely disappeared, when the addition of oxidative agent was combined with silencing of either PERK or NRF2.This further confirms the important role of NRF2 in blocking Cyclin D1 with stoichiometric inhibitors at early stage of oxidative stress.


Subject(s)
Cell Cycle Checkpoints/physiology , NF-E2-Related Factor 2/metabolism , Oxidative Stress/physiology , Cyclin D1/metabolism , HEK293 Cells , Humans , eIF-2 Kinase/metabolism
6.
Int J Mol Sci ; 18(1)2017 Jan 05.
Article in English | MEDLINE | ID: mdl-28067773

ABSTRACT

Accumulation of misfolded/unfolded proteins in the endoplasmic reticulum (ER) leads to the activation of three branches (Protein kinase (RNA)-like endoplasmic reticulum kinase [PERK], Inositol requiring protein 1 [IRE-1] and Activating trascription factor 6 [ATF6], respectively) of unfolded protein response (UPR). The primary role of UPR is to try to drive back the system to the former or a new homeostatic state by self-eating dependent autophagy, while excessive level of ER stress results in apoptotic cell death. Our study focuses on the role of PERK- and IRE-1-induced arms of UPR in life-or-death decision. Here we confirm that silencing of PERK extends autophagy-dependent survival, whereas the IRE-1-controlled apoptosis inducer is downregulated during ER stress. We also claim that the proper order of surviving and self-killing mechanisms is controlled by a positive feedback loop between PERK and IRE-1 branches. This regulatory network makes possible a smooth, continuous activation of autophagy with respect to ER stress, while the induction of apoptosis is irreversible and switch-like. Using our knowledge of molecular biological techniques and systems biological tools we give a qualitative description about the dynamical behavior of PERK- and IRE-1-controlled life-or-death decision. Our model claims that the two arms of UPR accomplish an altered upregulation of autophagy and apoptosis inducers during ER stress. Since ER stress is tightly connected to aging and age-related degenerative disorders, studying the signaling pathways of UPR and their role in maintaining ER proteostasis have medical importance.


Subject(s)
Endoplasmic Reticulum Stress/genetics , Signal Transduction/genetics , Systems Biology/methods , eIF-2 Kinase/genetics , Apoptosis/genetics , Autophagy/genetics , Blotting, Western , Cell Survival/genetics , Endoribonucleases/genetics , Endoribonucleases/metabolism , Feedback, Physiological , Gene Expression , HEK293 Cells , Homeostasis/genetics , Humans , Models, Biological , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , RNA Interference , Reverse Transcriptase Polymerase Chain Reaction , Unfolded Protein Response/genetics , eIF-2 Kinase/metabolism
7.
Biomed Res Int ; 2015: 319589, 2015.
Article in English | MEDLINE | ID: mdl-25984530

ABSTRACT

One of the most important tasks of a living organism is to maintain its genetic integrity with respect to stress. Endoplasmic reticulum (ER) has a crucial role in sensing cellular homeostasis by controlling metabolism, proteostasis, and several signaling processes. ER stressors can induce autophagy-dependent survival; however excessive level of stress results in apoptotic cell death. Although many molecular components of these networks have already been discovered, the analysis of the dynamical features of the regulatory network of life-or-death decision is still lacking. Our goal was to incorporate both theoretical and molecular biological techniques to explore the autophagy-apoptosis crosstalk under ER stress. Using various levels of different ER stressors we confirmed that the control network always generated an evidently detectable autophagy-dependent threshold for apoptosis activation. We explored the features of this threshold by introducing both autophagy activators and inhibitors, and transient treatment with excessive level of ER stressor was also performed. Our experimental data were also supported by a stochastic approach. Our analysis suggests that even if the switch-like characteristic of apoptosis activation is hardly seen on population level the double negative feedback loop between autophagy and apoptosis inducers introduces bistability in the control network.


Subject(s)
Apoptosis/drug effects , Autophagy/drug effects , Endoplasmic Reticulum Stress/drug effects , Cell Survival/drug effects , Computer Simulation , Dithiothreitol/pharmacology , HEK293 Cells , Hep G2 Cells , Humans , Models, Biological , Stochastic Processes , Time Factors , Tunicamycin/pharmacology
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