Dataset Information

Protease-independent control of parthanatos by HtrA2/Omi.

ABSTRACT: HtrA2/Omi is a mitochondrial serine protease with ascribed pro-apoptotic as well as pro-necroptotic functions. Here, we establish that HtrA2/Omi also controls parthanatos, a third modality of regulated cell death. Deletion of HtrA2/Omi protects cells from parthanatos while reconstitution with the protease restores the parthanatic death response. The effects of HtrA2/Omi on parthanatos are specific and cannot be recapitulated by manipulating other mitochondrial proteases such as PARL, LONP1 or PMPCA. HtrA2/Omi controls parthanatos in a manner mechanistically distinct from its action in apoptosis or necroptosis, i.e., not by cleaving cytosolic IAP proteins but rather exerting its effects without exiting mitochondria, and downstream of PARP-1, the first component of the parthanatic signaling cascade. Also, previously identified or candidate substrates of HtrA2/Omi such as PDXDC1, VPS4B or moesin are not cleaved and dispensable for parthanatos, whereas DBC-1 and stathmin are cleaved, and thus represent potential parthanatic downstream mediators of HtrA2/Omi. Moreover, mass-spectrometric screening for novel parthanatic substrates of HtrA2/Omi revealed that the induction of parthanatos does not cause a substantial proteolytic cleavage or major alterations in the abundance of mitochondrial proteins. Resolving these findings, reconstitution of HtrA2/Omi-deficient cells with a catalytically inactive HtrA2/Omi mutant restored their sensitivity against parthanatos to the same level as the protease-active HtrA2/Omi protein. Additionally, an inhibitor of HtrA2/Omi's protease activity did not confer protection against parthanatic cell death. Our results demonstrate that HtrA2/Omi controls parthanatos in a protease-independent manner, likely via novel, unanticipated functions as a scaffolding protein and an interaction with so far unknown mitochondrial proteins.

SUBMITTER: Weiß J

PROVIDER: S-EPMC10439076 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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Protease-independent control of parthanatos by HtrA2/Omi.

Weiß Jonas J Heib Michelle M Korn Thiemo T Hoyer Justus J Fuchslocher Chico Johaiber J Voigt Susann S Koudelka Tomas T Tholey Andreas A Adam Dieter D

Cellular and molecular life sciences : CMLS 20230818 9

HtrA2/Omi is a mitochondrial serine protease with ascribed pro-apoptotic as well as pro-necroptotic functions. Here, we establish that HtrA2/Omi also controls parthanatos, a third modality of regulated cell death. Deletion of HtrA2/Omi protects cells from parthanatos while reconstitution with the protease restores the parthanatic death response. The effects of HtrA2/Omi on parthanatos are specific and cannot be recapitulated by manipulating other mitochondrial proteases such as PARL, LONP1 or PM ...[more]

PMID: 37594630

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Project description:HtrA2(Omi) protease controls protein quality in mitochondria and plays a major role in apoptosis. Its HtrA2S306A mutant (with the catalytic serine routinely disabled for an X-ray study to avoid self-degradation) is a homotrimer whose subunits contain the serine protease domain (PD) and the regulatory PDZ domain. In the inactive state, a tight interdomain interface limits penetration of both PDZ-activating ligands and PD substrates into their respective target sites. We successfully crystalized HtrA2V226K/S306A, whose active counterpart HtrA2V226K has had higher proteolytic activity, suggesting higher propensity to opening the PD-PDZ interface than that of the wild type HtrA2. Yet, the crystal structure revealed the HtrA2V226K/S306A architecture typical of the inactive protein. To get a consistent interpretation of crystallographic data in the light of kinetic results, we employed molecular dynamics (MD). V325D inactivating mutant was used as a reference. Our simulations demonstrated that upon binding of a specific peptide ligand NH2-GWTMFWV-COOH, the PDZ domains open more dynamically in the wild type protease compared to the V226K mutant, whereas the movement is not observed in the V325D mutant. The movement relies on a PDZ vs. PD rotation which opens the PD-PDZ interface in a lid-like (budding flower-like in trimer) fashion. The noncovalent hinges A and B are provided by two clusters of interfacing residues, harboring V325D and V226K in the C- and N-terminal PD barrels, respectively. The opening of the subunit interfaces progresses in a sequential manner during the 50 ns MD simulation. In the systems without the ligand only minor PDZ shifts relative to PD are observed, but the interface does not open. Further activation-associated events, e.g. PDZ-L3 positional swap seen in any active HtrA protein (vs. HtrA2), were not observed. In summary, this study provides hints on the mechanism of activation of wtHtrA2, the dynamics of the inactive HtrA2V325D, but does not allow to explain an increased activity of HtrA2V226K.

Project description:The loss of mitochondrial function impairs intracellular energy production and potentially results in chronic liver disease. Increasing evidence suggests that mitochondrial dysfunction in hepatocytes contributes to the activation of hepatic stellate cells (HSCs), thereby resulting in hepatic fibrogenesis. High-temperature requirement protein A2 (HtrA2/Omi), a mitochondrial serine protease with various functions, is responsible for quality control in mitochondrial homeostasis. However, little information is available regarding its role in mitochondrial damage during the development of liver fibrosis. This study examined whether HtrA2/Omi regulates mitochondrial homeostasis in hepatocyte during the development of hepatic fibrogenesis. In this study, we demonstrated that HtrA2/Omi expression considerably decreased in liver tissues from the CCl4-induced liver fibrotic mice model and from patients with liver cirrhosis. Knockdown of HtrA2/Omi in hepatocytes induced the accumulation of damaged mitochondria and provoked mitochondrial reactive oxygen species (mtROS) stress. We further show that the damaged mtDNA isolated from HtrA2/Omi-deficient hepatocytes as a form of damage-associated molecular patterns can induce HSCs activation. Moreover, we found that motor neuron degeneration 2-mutant mice harboring the missense mutation Ser276Cys in the protease domain of HtrA2/Omi displayed altered mitochondrial morphology and function, which increased oxidative stress and promoted liver fibrosis. Conversely, the overexpression of HtrA2/Omi via hydrodynamics-based gene transfer led to the antifibrotic effects in CCl4-induced liver fibrosis mice model through decreasing collagen accumulation and enhancing anti-oxidative activity by modulating mitochondrial homeostasis in the liver. These results suggest that suppressing HtrA2/Omi expression promotes hepatic fibrogenesis via modulating mtROS generation, and these novel mechanistic insights involving the regulation of mitochondrial homeostasis by HtrA2/Omi may be of importance for developing new therapeutic strategies for hepatic fibrosis.

Dataset Information

Protease-independent control of parthanatos by HtrA2/Omi.

Publications

Protease-independent control of parthanatos by HtrA2/Omi.

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