RESUMEN
PURPOSE: HtrA1, HtrA2, HtrA3 and HtrA4 appear to be involved in the development of pathologies such as cancer. This systematic review reports the results of a literature search performed to compare the expression of HtrA family genes and proteins in cancer versus non-cancer tissues and cell lines, assess relationships between HtrA expression and cancer clinical features in cancer, and analyse the molecular mechanism, by which HtrA family affects cancer. METHODS: The literature search was conducted according to the PRISMA statement among four databases (PubMed, Web of Science, Embase and Scopus). RESULTS: A total of 38 articles met the inclusion criteria and involved the expression of HtrA family members and concerned the effect of HtrA expression on cancer and metastasis development or on the factor that influences it. Additionally, 31 reports were retrieved manually. Most articles highlighted that HtrA1 and HtrA3 exhibited tumour suppressor activity, while HtrA2 was associated with tumour growth and metastasis. There were too few studies to clearly define the role of the HtrA4 protease in tumours. CONCLUSION: Although the expression of serine proteases of the HtrA family was dependent on tumour type, stage and the presence of metastases, most articles indicated that HtrA1 and HtrA3 expression in tumours was downregulated compared with healthy tissue or cell lines. The expression of HtrA2 was completely study dependent. The limited number of studies on HtrA4 expression made it impossible to draw conclusions about differences in expression between healthy and tumour tissue. The conclusions drawn from the study suggest that HtrA1 and HtrA3 act as tumour suppressors.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Serina Peptidasa A1 que Requiere Temperaturas Altas , Serina Peptidasa A2 que Requiere Temperaturas Altas , Neoplasias , Serina Endopeptidasas , Humanos , Neoplasias/genética , Neoplasias/patología , Serina Peptidasa A1 que Requiere Temperaturas Altas/genética , Serina Peptidasa A1 que Requiere Temperaturas Altas/metabolismo , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismoRESUMEN
The mitochondrial serine protease HtrA2 is a human homolog of the Escherichia coli Deg-proteins exhibiting chaperone and proteolytic roles. HtrA2 is involved in both apoptotic regulation via its ability to degrade inhibitor-of-apoptosis proteins (IAPs), as well as in cellular maintenance as part of the cellular protein quality control machinery, by preventing the possible toxic accumulation of aggregated proteins. In this study, we use advanced solution NMR spectroscopy methods combined with biophysical characterization and biochemical assays to elucidate the crucial role of the substrate recognizing PDZ domain. This domain regulates the protease activity of HtrA2 by triggering an intricate allosteric network involving the regulatory loops of the protease domain. We further show that divalent metal ions can both positively and negatively modulate the activity of HtrA2, leading to a refined model of HtrA2 regulation within the apoptotic pathway.
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Apoptosis , Serina Peptidasa A2 que Requiere Temperaturas Altas , Dominios PDZ , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Humanos , Regulación Alostérica , Especificidad por Sustrato , Mitocondrias/metabolismo , Modelos Moleculares , Espectroscopía de Resonancia MagnéticaRESUMEN
Aggregation and misfolding of α-Synuclein (α-Syn), a causative agent for Parkinson's disease (PD), and oxidative stress are tightly implicated in the pathogenesis of PD. Although more than 20 genes including HtrA2 have been identified as causative genes for PD, the molecular mechanisms underlying the pathophysiological functions between HtrA2 and α-Syn in the pathogenesis of PD remain unclear. This study shows that HtrA2 serine protease selectively recognizes and interacts with the NAC region of α-Syn. Interestingly, we found that HtrA2 causes proteolysis of α-Syn to prevent mitochondrial accumulation of α-Syn, thereby inhibiting the production of reactive oxygen species (ROS) in the mitochondria. We have further demonstrated that HtrA2 knockdown promotes α-Syn-mediated mitochondrial ROS production, thereby activating microglial cells. This study is the first to demonstrate that the HtrA2/α-Syn cellular partner may play a crucial role in the pathogenesis of PD and provide new insights into the pathological processes and effective therapeutic strategies for PD.
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Enfermedad de Parkinson , alfa-Sinucleína , Humanos , alfa-Sinucleína/genética , Especies Reactivas de Oxígeno , Microglía/patología , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Mitocondrias/patologíaRESUMEN
The premature death and degeneration of striatal neurons are typical hallmarks of HtrA2-inactivated motor neuron degeneration 2 (mnd2) mice. Although HtrA2 has been extensively studied in relation to the regulation of apoptosis using mnd2 mice, little is known about the other physiological functions of HtrA2. In this study, we found that the skin color of wild-type (WT) and mnd2 mice was black and pink on postnatal day 32. Using histological and molecular assays (i.e., assessing the activation of MAPK and expression patterns of PCNA), we demonstrated that this differential skin color change is consistent with the delay in the telogen - to - anagen phase of the hair cycle in mnd2 mice. We also examined adipocytes in the subcutaneous skin layer, finding that HtrA2 inactivation leads to the growth retardation of adipocytes, thereby delaying the hair cycle of mnd2 mice. Collectively, these findings show for the first time that HtrA2 plays an essential role in regulating the adipogenesis-associated hair cycle.
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Proteínas Mitocondriales , Serina Endopeptidasas , Animales , Ratones , Apoptosis , Cabello/metabolismo , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Proteínas Mitocondriales/metabolismo , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismoRESUMEN
Members of the HtrA family of serine proteases are known to play roles in mitochondrial homeostasis as well as in programmed cell death. Mitochondrial homeostasis and metabolism are crucial for the survival and propagation of the malaria parasite within the host. Here we have functionally characterized a Plasmodium falciparum HtrA2 (PfHtrA2) protein, which harbours trypsin-like protease activity that can be inhibited by its specific inhibitor, ucf-101. A transgenic parasite line was generated, using the HA-glmS C-terminal tagging approach, for localization as well as for inducible knock-down of PfHtrA2. The PfHtrA2 was localized in the parasite mitochondrion during the asexual life cycle. Genetic ablation of PfHtrA2 caused significant parasite growth inhibition, decreased replication of mtDNA, increased mitochondrial ROS production, caused mitochondrial fission/fragmentation, and hindered parasite development. However, the ucf-101 treatment did not affect the parasite growth, suggesting the non-protease/chaperone role of PfHtrA2 in the parasite. Under cellular stress conditions, inhibition of PfHtrA2 by ucf-101 reduced activation of the caspase-like protease as well as parasite cell death, suggesting the involvement of protease activity of PfHtrA2 in apoptosis-like cell death in the parasite. Under these cellular stress conditions, the PfHtrA2 gets processed but remains localized in the mitochondrion, suggesting that it acts within the mitochondrion by cleaving intra-mitochondrial substrate(s). This was further supported by trans-expression of PfHtrA2 protease domain in the parasite cytosol, which was unable to induce any cell death in the parasite. Overall, we show the specific roles of PfHtrA2 in maintaining mitochondrial homeostasis as well as in regulating stress-induced cell death.
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Malaria , Parásitos , Animales , Humanos , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Parásitos/metabolismo , Proteínas Mitocondriales/metabolismo , Mitocondrias/metabolismo , Apoptosis , Muerte Celular , Homeostasis , Malaria/metabolismoRESUMEN
Skeletal muscles require muscle satellite cell (MuSC) differentiation to facilitate the replenishment and repair of muscle fibers. A key step in this process is called myogenic differentiation. The differentiation ability of MuSCs decreases with age and can result in sarcopenia. Although mitochondria have been reported to be involved in myogenic differentiation by promoting a bioenergetic remodeling, little is known about the interplay of mitochondrial proteostasis and myogenic differentiation. High-temperature-requirement protein A2 (HtrA2/Omi) is a protease that regulates proteostasis in the mitochondrial intermembrane space (IMS). Mice deficient in HtrA2 protease activity show a distinct phenotype of sarcopenia. To investigate the role of IMS proteostasis during myogenic differentiation, we treated C2C12 myoblasts with UCF101, a specific inhibitor of HtrA2 during differentiation process. A key step in this process is called myogenic differentiation. The differentiation ability of MuSCs decreases with age and can result in sarcopenia. Further, CHOP, p-eIF2α, and other mitochondrial unfolded protein response (UPRmt)-related proteins are upregulated. Therefore, we suggest that imbalance of mitochondrial IMS proteostasis acts via a retrograde signaling pathway to inhibit myogenic differentiation via the UPRmt pathway. These novel mechanistic insights may have implications for the development of new strategies for the treatment of sarcopenia.
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Serina Peptidasa A2 que Requiere Temperaturas Altas , Proteínas Mitocondriales , Sarcopenia , Animales , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Ratones , Proteínas Mitocondriales/metabolismo , Péptido HidrolasasRESUMEN
Radiation therapy can induce cellular senescence in cancer cells, leading to short-term tumor growth arrest but increased long-term recurrence. To better understand the molecular mechanisms involved, we developed a model of radiation-induced senescence in cultured cancer cells. The irradiated cells exhibited a typical senescent phenotype, including upregulation of p53 and its main target, p21, followed by a sustained reduction in cellular proliferation, changes in cell size and cytoskeleton organization, and senescence-associated beta-galactosidase activity. Mass spectrometry-based proteomic profiling of the senescent cells indicated downregulation of proteins involved in cell cycle progression and DNA repair, and upregulation of proteins associated with malignancy. A functional siRNA screen using a cell death-related library identified mitochondrial serine protease HtrA2 as being necessary for sustained growth arrest of the senescent cells. In search of direct HtrA2 substrates following radiation, we determined that HtrA2 cleaves the intermediate filament protein vimentin, affecting its cytoplasmic organization. Ectopic expression of active cytosolic HtrA2 resulted in similar changes to vimentin filament assembly. Thus, HtrA2 is involved in the cytoskeletal reorganization that accompanies radiation-induced senescence and the continuous maintenance of proliferation arrest.
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Senescencia Celular , Serina Peptidasa A2 que Requiere Temperaturas Altas , Neoplasias , Proteómica , Apoptosis , Senescencia Celular/fisiología , Senescencia Celular/efectos de la radiación , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Humanos , Proteínas Mitocondriales/metabolismo , Neoplasias/genética , Neoplasias/radioterapia , Células Tumorales Cultivadas , Vimentina/metabolismoRESUMEN
High temperature requirement A2 (HtrA2) contributes to regulating mitochondrial quality control and maintaining the balance between the death and survival of cells and living organisms. However, the molecular mechanism of HtrA2 in physiological and pathophysiological processes remains unclear. HtrA2 exhibits multifaceted characteristics according to the expression levels and acts opposite functions depending on its subcellular localization. Thus, innovative technologies and systems that can be freely manipulated at the quantitative, biochemical, molecular and cellular levels are needed to address not only the challenges faced by HtrA2 research but also the general obstacles to protein research. Here, we are the first to identify zebrafish HtrA2 (zHtrA2) as the true ortholog of human HtrA2 (hHtrA2), by in silico sequence analysis of genomic DNA and molecular biological techniques, which is highly conserved structurally and functionally as a serine protease and cell death regulator. The zHtrA2 protein is primarily localized in the mitochondria, where alanine-exposed mature zHtrA2 ((A)-zHtrA2) is generated by removing 111 residues at the N-terminus of pro-zHtrA2. The (A)-zHtrA2 released from the mitochondria into the cytosol induces the caspase cascade by binding to and inhibiting hXIAP, a cognate partner of hHtrA2. Notably, zHtrA2 has well conserved properties of serine protease that specifically cleaves hParkin, a cognate substrate of hHtrA2. Interestingly, cytosolic (M)-zHtrA2, which does not bind hXIAP, induces atypical cell death in a serine protease-dependent manner, as occurs in hHtrA2. Thus, the zebrafish-zHtrA2 system can be used to clarify the crucial role of HtrA2 in maintaining the survival of living organisms and provide an opportunity to develop novel therapeutics for HtrA2-associated diseases, such as neurodegenerative diseases and cancer, which are caused by dysregulation of HtrA2.
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Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Homeostasis , Mitocondrias/genética , Animales , Caspasas/metabolismo , Muerte Celular , Genes Mitocondriales , Células HEK293 , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Humanos , Mitocondrias/metabolismo , Proteínas Mitocondriales/genética , Pez Cebra , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
BACKGROUND: mRNAs have been shown to be critical biomarkers or therapeutic targets for human diseases. However, only a few of them have been studied as blood-based biomarkers for gastric carcinoma (GC) detection. METHODS: mRNA expression profiles for GC were screened using plasma samples from 10 GC patients with different TNM stages and 5 healthy individuals as controls. One candidate tumor-related mRNA named HTRA2 was then evaluated in GC samples with quantitative real-time polymerase chain reaction (qRT-PCR). TCGAportal, UALCAN, and TISCH database were used to explore the function of HTRA2 in GC. Finally, the effect generated by HTRA2 expression on cell proliferating, invading, and migrating processes was assessed in vitro with knockdown and over-expression strategies. RESULTS: HTRA2 displayed noticeable increase inside GC plasma compared with control cases. Higher expression of HTRA2 displayed a correlation to higher clinicopathological stage and worse prognosis. HTRA2 knocking down down-regulated GC cells' proliferating, invading, and migrating states, while HTRA2 over-expression exerted the inconsistent influence. HTRA2 protein, which may interact with PINK1, PARL, and CYCS, was mainly located in the mitochondria of cells and primarily involved cellular response and metabolic signaling pathway. Immune factors may interact with HTRA2 in GC, and HTRA2 was found noticeably linked with immunosuppressor such as CD274, IDO1, and TIGIT. CONCLUSION: One plasma HTRA2 can be an emerging diagnosis-related biomarker to achieve GC detecting process, but the particular regulatory effect still needs to be further explored.
Asunto(s)
Ácidos Nucleicos Libres de Células/sangre , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Neoplasias Gástricas/genética , Biomarcadores de Tumor/genética , Estudios de Casos y Controles , Regulación Neoplásica de la Expresión Génica , Estudio de Asociación del Genoma Completo , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , ARN Mensajero/sangre , Transducción de Señal/genética , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/patologíaRESUMEN
The human high-temperature requirement A2 (HtrA2) mitochondrial protease is critical for cellular proteostasis, with mutations in this enzyme closely associated with the onset of neurodegenerative disorders. HtrA2 forms a homotrimeric structure, with each subunit composed of protease and PDZ (PSD-95, DLG, ZO-1) domains. Although we had previously shown that successive ligand binding occurs with increasing affinity, and it has been suggested that allostery plays a role in regulating catalysis, the molecular details of how this occurs have not been established. Here, we use cysteine-based chemistry to generate subunits in different conformational states along with a protomer mixing strategy, biochemical assays, and methyl-transverse relaxation optimized spectroscopy-based NMR studies to understand the role of interprotomer allostery in regulating HtrA2 function. We show that substrate binding to a PDZ domain of one protomer increases millisecond-to-microsecond timescale dynamics in neighboring subunits that prime them for binding substrate molecules. Only when all three PDZ-binding sites are substrate bound can the enzyme transition into an active conformation that involves significant structural rearrangements of the protease domains. Our results thus explain why when one (or more) of the protomers is fixed in a ligand-binding-incompetent conformation or contains the inactivating S276C mutation that is causative for a neurodegenerative phenotype in mouse models of Parkinson's disease, transition to an active state cannot be formed. In this manner, wild-type HtrA2 is only active when substrate concentrations are high and therefore toxic and unregulated proteolysis of nonsubstrate proteins can be suppressed.
Asunto(s)
Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Mitocondrias/metabolismo , Mutación , Dominios PDZ , Enfermedad de Parkinson/patología , Regiones Promotoras Genéticas , Animales , Dominio Catalítico , Serina Peptidasa A2 que Requiere Temperaturas Altas/química , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Humanos , Ratones , Mitocondrias/genética , Modelos Moleculares , Enfermedad de Parkinson/etiología , Conformación Proteica , Proteolisis , Relación Estructura-ActividadRESUMEN
HtrA2, a proapoptotic mitochondrial serine protease, promotes cellular protection against oxidative damage. Literature reports show positive correlation between loss of HtrA2 protease activity and Parkinson's Disease (PD) susceptibility. Homozygous loss-of-function mutations in murine-HtrA2, and when they rarely occur in humans result in severe neurodegeneration and infantile death. Here, we report a novel heterozygous pathogenic HTRA2 variant, c.725C > T (p.T242M) in Indian PD patients. Although, this mutation exhibits no significant conformational changes compared to the wild-type, functional studies with HtrA2-T242M transfected neurons reveal common features of PD pathogenesis such as dysfunction, altered morphology and mitochondrial membrane depolarization. Despite exhibiting two-fold decrease in enzyme activity, observation of excessive cell-death due to over-expression of the mutant has been correlated with it being constitutively active. This interesting behavioral anomaly has been attributed to the loss of phosphorylation-mediated regulatory checkpoint at the T242M mutation site that is otherwise controlled by glycogen synthase kinase-3ß (GSK-3ß). This study, with seamless amalgamation of biophysical and biomedical research unravels a mechanistic pathway of HtrA2 regulation and delineates its biological role in PD. Therefore, this investigation will not only prove beneficial toward devising therapeutic strategies against HtrA2-associated diseases mediated by GSK-3ß but also suggest new avenues for treatment of Parkinsonian phenotype.
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Apoptosis/genética , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Mutación con Pérdida de Función , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/metabolismo , Fenotipo , Adulto , Estudios de Casos y Controles , Línea Celular Tumoral , Femenino , Glucógeno Sintasa Quinasa 3 beta/genética , Células HEK293 , Heterocigoto , Serina Peptidasa A2 que Requiere Temperaturas Altas/química , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Humanos , India/epidemiología , Masculino , Persona de Mediana Edad , Mitocondrias/metabolismo , Neuronas/metabolismo , Enfermedad de Parkinson/epidemiología , Fosforilación/genética , Polimorfismo de Nucleótido Simple , Estructura Secundaria de Proteína , Transfección , Adulto JovenRESUMEN
BACKGROUND: Aging, noise, infection, and ototoxic drugs are the major causes of human acquired sensorineural hearing loss, but treatment options are limited. CRISPR/Cas9 technology has tremendous potential to become a new therapeutic modality for acquired non-inherited sensorineural hearing loss. Here, we develop CRISPR/Cas9 strategies to prevent aminoglycoside-induced deafness, a common type of acquired non-inherited sensorineural hearing loss, via disrupting the Htra2 gene in the inner ear which is involved in apoptosis but has not been investigated in cochlear hair cell protection. RESULTS: The results indicate that adeno-associated virus (AAV)-mediated delivery of CRISPR/SpCas9 system ameliorates neomycin-induced apoptosis, promotes hair cell survival, and significantly improves hearing function in neomycin-treated mice. The protective effect of the AAV-CRISPR/Cas9 system in vivo is sustained up to 8 weeks after neomycin exposure. For more efficient delivery of the whole CRISPR/Cas9 system, we also explore the AAV-CRISPR/SaCas9 system to prevent neomycin-induced deafness. The in vivo editing efficiency of the SaCas9 system is 1.73% on average. We observed significant improvement in auditory brainstem response thresholds in the injected ears compared with the non-injected ears. At 4 weeks after neomycin exposure, the protective effect of the AAV-CRISPR/SaCas9 system is still obvious, with the improvement in auditory brainstem response threshold up to 50 dB at 8 kHz. CONCLUSIONS: These findings demonstrate the safe and effective prevention of aminoglycoside-induced deafness via Htra2 gene editing and support further development of the CRISPR/Cas9 technology in the treatment of non-inherited hearing loss as well as other non-inherited diseases.
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Sistemas CRISPR-Cas , Edición Génica , Pérdida Auditiva Sensorineural/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Animales , Línea Celular , Supervivencia Celular/genética , Dependovirus/genética , Modelos Animales de Enfermedad , Expresión Génica , Marcación de Gen , Ingeniería Genética , Terapia Genética/métodos , Vectores Genéticos/administración & dosificación , Vectores Genéticos/genética , Células Ciliadas Auditivas/metabolismo , Pérdida Auditiva Sensorineural/inducido químicamente , Pérdida Auditiva Sensorineural/etiología , Pérdida Auditiva Sensorineural/terapia , Pruebas Auditivas , Ratones , ARN Guía de Kinetoplastida , Transducción Genética , Transgenes , Resultado del TratamientoRESUMEN
Serine protease Omi/HtrA2, a member of the HtrA family, is closely related to the maintenance of mitochondrial integrity and participates in apoptosis but its role in cerebral ischemia/reperfusion (I/R) injury and cellular oxidative stress response remains unclear. In this study, we found that I/R injury resulted in a time-dependent increase in Omi/HtrA2 expression in rat brain tissue. Inhibition of Omi/HtrA2 significantly inhibited XIAP cleavage in H2O2-induced PC12 cells. In addition, inhibition of Omi/HtrA2 significantly inhibited the up-regulation of mitochondrial stress proteins CHOP and ClpP, significantly reduced mitochondrial aggregation, and attenuated the decline of mitochondrial ΔΨm in PC12 cells. Studies show that there is a physical interaction between Omi/HtrA2 and OPA1. We found that Omi/HtrA2 and OPA1 are closely related to the oxidative stress mitochondrial response in PC12 cells. The current study has demonstrated that Omi/HtrA2 is upregulated in brain I/R injury in vivo and is implicated in mitochondrial response to oxidative stress in vitro by regulating mitochondrial stress proteins CHOP and CLpP and by interacting with mitochondrial cristae remodeling protein OPA1. These findings suggest that Omi/HtrA2 could be a candidate molecular target in diseases that involve oxidative stress such as in I/R injury. Abbreviation: ATP: Adenosine tripHospHate; Bax: BCL2-Associated X; Bcl-2: B-cell lympHoma-2; BSA: Albumin from bovine serum; DMEM: Dulbecco's Minimum Essential Medium; DMSO: Dimethyl sulfoxide; HSP60: Heat shock protein60, 70; L-OPA1: Long forms of OPA1; Omi/HtrA2: high-temperature-regulated A2; MCAO: Middle cerebral artery occlusion; OPA1: Optic AtropHy; PBS: PHospHate buffered saline; PMSF: pHenylmethyl sulfonylfluoride; ROS: reactive oxygen species; SDS: Sodium dodecyl sulfate; S-OPA1: Short forms of OPA1; TTC: TripHenyltetrazalium chloride; XIAP: X-linked inhibitor apoptosis protein.
Asunto(s)
GTP Fosfohidrolasas/metabolismo , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Proteínas Mitocondriales/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Factor de Transcripción CHOP/metabolismo , Animales , Apoptosis/fisiología , GTP Fosfohidrolasas/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Peróxido de Hidrógeno/farmacología , Masculino , Mitocondrias/metabolismo , Dinámicas Mitocondriales/genética , Dinámicas Mitocondriales/fisiología , Proteínas Mitocondriales/genética , Estrés Oxidativo/fisiología , Células PC12 , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Factor de Transcripción CHOP/genéticaRESUMEN
HtrA proteases regulate cellular homeostasis and cell death. Their dysfunctions have been correlated with oncogenesis and response to therapeutic treatment. We investigated the relation between HtrA1-3 expression and clinicopathological, and survival data, as well as the microsatellite status of tumors. Sixty-five colorectal cancer patients were included in the study. The expression of HTRA1-3 was estimated at the mRNA and protein levels by quantitative PCR and immunoblotting. Microsatellite status was determined by high-resolution-melting PCR. We found that the HTRA1 mRNA level was higher in colorectal cancer tissue as compared to the unchanged mucosa, specifically in primary lesions of metastasizing cancer. The levels of HtrA1 and HtrA2 proteins were reduced in tumor tissue when compared to unchanged mucosa, specifically in primary lesions of metastasizing disease. Moreover, a decrease in HTRA1 and HTRA2 transcripts' levels in cancers with a high level of microsatellite instability compared to microsatellite stable ones has been observed. A low level of HtrA1 or/and HtrA2 in cancer tissue correlated with poorer patient survival. The expression of HTRA1 and HTRA2 changes during colorectal carcinogenesis and microsatellite instability may be, at least partially, associated with these changes. The alterations in the HTRA1/2 genes' expression are connected with metastatic potential of colorectal cancer and may affect patient survival.
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Neoplasias Colorrectales/genética , Regulación Neoplásica de la Expresión Génica , Serina Peptidasa A1 que Requiere Temperaturas Altas/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Inestabilidad de Microsatélites , Serina Endopeptidasas/genética , Adulto , Anciano , Supervivencia Celular , Neoplasias Colorrectales/mortalidad , Femenino , Humanos , Masculino , Repeticiones de Microsatélite , Persona de Mediana Edad , Metástasis de la Neoplasia , Isoformas de ProteínasRESUMEN
Mitochondrial dysfunction has long been implicated in the neurodegenerative disorder Parkinson's disease (PD); however, it is unclear how mitochondrial impairment and α-synuclein pathology are coupled. Using specific mitochondrial inhibitors, EM analysis, and biochemical assays, we report here that intramitochondrial protein homeostasis plays a major role in α-synuclein aggregation. We found that interference with intramitochondrial proteases, such as HtrA2 and Lon protease, and mitochondrial protein import significantly aggravates α-synuclein seeding. In contrast, direct inhibition of mitochondrial complex I, an increase in intracellular calcium concentration, or formation of reactive oxygen species, all of which have been associated with mitochondrial stress, did not affect α-synuclein pathology. We further demonstrate that similar mechanisms are involved in amyloid-ß 1-42 (Aß42) aggregation. Our results suggest that, in addition to other protein quality control pathways, such as the ubiquitin-proteasome system, mitochondria per se can influence protein homeostasis of cytosolic aggregation-prone proteins. We propose that approaches that seek to maintain mitochondrial fitness, rather than target downstream mitochondrial dysfunction, may aid in the search for therapeutic strategies to manage PD and related neuropathologies.
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Péptidos beta-Amiloides/metabolismo , Mitocondrias/metabolismo , Enfermedad de Parkinson/metabolismo , Fragmentos de Péptidos/metabolismo , Proteostasis , alfa-Sinucleína/metabolismo , Péptidos beta-Amiloides/genética , Animales , Línea Celular Tumoral , Femenino , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Humanos , Mitocondrias/genética , Mitocondrias/patología , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Fragmentos de Péptidos/genética , Ratas , Ratas Sprague-Dawley , Factores de Empalme Serina-Arginina/genética , Factores de Empalme Serina-Arginina/metabolismo , alfa-Sinucleína/genéticaRESUMEN
Cellular homeostasis requires tight coordination between nucleus and mitochondria, organelles that each possesses their own genomes. Disrupted mitonuclear communication has been found to be implicated in many aging processes. However, little is known about mitonuclear signaling regulator in sarcopenia which is a major contributor to the risk of poor health-related quality of life, disability, and premature death in older people. High-temperature requirement protein A2 (HtrA2/Omi) is a mitochondrial protease and plays an important role in mitochondrial proteostasis. HtrA2mnd2(-/-) mice harboring protease-deficient HtrA2/Omi Ser276Cys missense mutants exhibit premature aging phenotype. Additionally, HtrA2/Omi has been established as a signaling regulator in nervous system and tumors. We therefore asked whether HtrA2/Omi participates in mitonuclear signaling regulation in muscle degeneration. Using motor functional, histological, and molecular biological methods, we characterized the phenotype of HtrA2mnd2(-/-) muscle. Furthermore, we isolated the gastrocnemius muscle of HtrA2mnd2(-/-) mice and determined expression of genes in mitochondrial unfolded protein response (UPRmt ), mitohormesis, electron transport chain (ETC), and mitochondrial biogenesis. Here, we showed that HtrA2/Omi protease deficiency induced denervation-independent skeletal muscle degeneration with sarcopenia phenotypes. Despite mitochondrial hypofunction, upregulation of UPRmt and mitohormesis-related genes and elevated total reactive oxygen species (ROS) production were not observed in HtrA2mnd2(-/-) mice, contrary to previous assumptions that loss of protease activity of HtrA2/Omi would lead to mitochondrial dysfunction as a result of proteostasis disturbance and ROS burst. Instead, we showed that HtrA2/Omi protease deficiency results in different changes between the expression of nuclear DNA- and mitochondrial DNA-encoded ETC subunits, which is in consistent with their transcription factors, nuclear respiratory factors 1 and 2, and coactivator peroxisome proliferator-activated receptor γ coactivator 1α. These results reveal that loss of HtrA2/Omi protease activity induces mitonuclear imbalance via differential regulation of mitochondrial biogenesis in sarcopenia. The novel mechanistic insights may be of importance in developing new therapeutic strategies for sarcopenia.
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Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Biogénesis de Organelos , Sarcopenia/genética , Anciano , Anciano de 80 o más Años , Animales , Núcleo Celular/genética , Proteínas del Complejo de Cadena de Transporte de Electrón/genética , Homeostasis/genética , Humanos , Masculino , Ratones , Ratones Noqueados , Mitocondrias/genética , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Sarcopenia/metabolismo , Sarcopenia/patología , Temperatura , Respuesta de Proteína Desplegada/genéticaRESUMEN
Although the malfunction of HtrA2/Omi leads to Parkinson's disease (PD), the underlying mechanism has remained unknown. Here, we showed that HtrA2/Omi specifically removed oligomeric α-Syn but not monomeric α-Syn to protect oligomeric α-Syn-induced neurodegeneration. Experiments using mnd2 mice indicated that HtrA2/Omi degraded oligomeric α-Syn specifically without affecting monomers. Transgenic Drosophila melanogaster experiments of the co-expression α-Syn and HtrA2/Omi and expression of genes individually also confirmed that pan-neuronal expression of HtrA2/Omi completely rescued Parkinsonism in the α-Syn-induced PD Drosophila model by specifically removing oligomeric α-Syn. HtrA2/Omi maintained the health and integrity of the brain and extended the life span of transgenic flies. Because HtrA2/Omi specifically degraded oligomeric α-Syn, co-expression of HtrA2/Omi and α-Syn in Drosophila eye maintained a healthy retina, while the expression of α-Syn induced retinal degeneration. This work showed that the bacterial function of HtrA to degrade toxic misfolded proteins is evolutionarily conserved in mammalian brains as HtrA2/Omi.
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Encéfalo/metabolismo , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Neuronas/metabolismo , Enfermedad de Parkinson/prevención & control , alfa-Sinucleína/metabolismo , Animales , Animales Modificados Genéticamente , Encéfalo/patología , Modelos Animales de Enfermedad , Drosophila melanogaster , Femenino , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/patología , Enfermedad de Parkinson/metabolismo , Enfermedad de Parkinson/patología , alfa-Sinucleína/genéticaRESUMEN
INTRODUCTION: Essential tremor (ET) is one of the most prevalent movement disorders. The genetic etiology of ET has not been well defined although a significant proportion (≥50%) are familial cases. Linkage analysis and genome-wide association studies (GWASs) have identified several risk variants. In recent years, whole-exome sequencing of ET has revealed several specific causal variants in FUS (p.Q290X), HTRA2 (p.G399S), and TENM4 (c.4324 G>A, c.4100C>A, and c.3412G>A) genes. OBJECTIVE: To investigate the genetic contribution of these three genes to ET, the protein-coding sequences of FUS, HTRA2, and TENM4 were analyzed in a total of 238 ET patients and 272 controls from eastern China using direct Sanger sequencing. RESULTS: We identified two synonymous coding single nucleotide polymorphisms (SNPs), rs741810 and rs1052352 in FUS, and three previously reported synonymous SNPs, rs11237621, rs689369, and rs2277277 in TENM4. No nonsynonymous exonic variants were identified in these subjects. We found that the frequency of the rs1052352C allele was significantly higher (P = .001) in the ET group than in the control group. CONCLUSION: Overall, our findings suggest that rs1052352 of FUS might contribute to ET risk in Chinese population.
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Pueblo Asiatico/genética , Temblor Esencial/genética , Pruebas Genéticas/métodos , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Glicoproteínas de Membrana/genética , Proteína FUS de Unión a ARN/genética , Adolescente , Adulto , Anciano , Niño , Temblor Esencial/diagnóstico , Temblor Esencial/epidemiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple/genética , Adulto JovenRESUMEN
Acetaldehyde dehydrogenase 2 (ALDH2) is an enzyme involved in redox homeostasis as well as the detoxification process in alcohol metabolism. Nearly 8% of the world's population have an inactivating mutation in the ALDH2 gene. However, the expression patterns and specific functions of ALDH2 in skeletal muscles are still unclear. Herein, we report that ALDH2 is expressed in skeletal muscle and is localized to the mitochondrial fraction. Oxidative muscles had a higher amount of ALDH2 protein than glycolytic muscles. We next comprehensively investigated whether ALDH2 knockout in mice induces mitochondrial adaptations in gastrocnemius muscle (for example, content, enzymatic activity, respiratory function, supercomplex formation, and functional networking). We found that ALDH2 deficiency resulted in partial mitochondrial dysfunction in gastrocnemius muscle because it increased mitochondrial reactive oxygen species (ROS) emission (2',7'-dichlorofluorescein and MitoSOX oxidation rate during respiration) and the frequency of regional mitochondrial depolarization. Moreover, we determined whether ALDH2 deficiency and the related mitochondrial dysfunction trigger mitochondrial stress and quality control responses in gastrocnemius muscle (for example, mitophagy markers, dynamics, and the unfolded protein response). We found that ALDH2 deficiency upregulated the mitochondrial serine protease Omi/HtrA2 (a marker of the activation of a branch of the mitochondrial unfolded protein response). In summary, ALDH2 deficiency leads to greater mitochondrial ROS production, but homeostasis can be maintained via an appropriate stress response.
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Aldehído Deshidrogenasa Mitocondrial/metabolismo , Genotipo , Serina Peptidasa A2 que Requiere Temperaturas Altas/metabolismo , Mitocondrias/metabolismo , Músculo Esquelético/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Aldehído Deshidrogenasa Mitocondrial/genética , Animales , Regulación de la Expresión Génica , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Consumo de OxígenoRESUMEN
The main pathological hallmark of Parkinson's disease (PD) is the presence of Lewy bodies, which mainly consist of aggregated α-synuclein. Based on the neurotoxicity of oligomeric α-synuclein and its significance in the aetiology of PD, there has been decades of effort to elucidate an enzyme specifically degrading oligomeric α-synuclein. Here we report an enzyme, Omi, which specifically recognizes and precisely degrades oligomeric α-synuclein but not monomeric α-synuclein. After enzymatic and functional analyses of Omi in in vitro, we developed an in vivo assay system of dual gene interaction in Drosophila to investigate further the etiological role of Omi in PD. Pan-neuronal expression of Omi rescued Parkinsonism in a Drosophila model of PD, while Knockout of Omi exacerbated Parkinsonism. Expression of Omi counteracted the α-synuclein-induced retinal degeneration, providing additional evidence for Omi's protective role oligomeric α-synuclein. This work reports identification of the catabolic pathway of oligomeric α-synuclein as well as showing how Omi functions as the key molecule in the recognition and degradation of toxic oligomeric α-synuclein, a possible cause of neurodegeneration in PD, without affecting monomeric α-synuclein which is a native essential molecule for the normal function of neurons.