Research progress in the role of E3 ubiquitin ligases in viral infection / 中华微生物学和免疫学杂志

Ubiquitination modifications are a kind of post-translational modifications of proteins widely found in eukaryotes and involved in a variety of biological activities. E3 ubiquitin ligases are an important component of the ubiquitin system, with the function of specific recognition of substrate proteins and mediation of different types of ubiquitination modifications. They can regulate the function and life time of substrate proteins. Recent studies have shown that E3 ubiquitin ligases are widely involved in the regulation of the host innate immune response and can directly or indirectly influence viral infection. Moreover, viruses are able to encode or hijack E3 ubiquitin ligases in their long-term evolution, allowing them to play an important role in viral infection and replication cycle. This paper reviewed the progress in the mechanisms of E3 ubiquitin ligases in innate immune responses and viral infection in recent years.

The role of E3 ubiquitin ligases in bone homeostasis and related diseases

The ubiquitin-proteasome system (UPS) dedicates to degrade intracellular proteins to modulate demic homeostasis and functions of organisms. These enzymatic cascades mark and modifies target proteins diversly through covalently binding ubiquitin molecules. In the UPS, E3 ubiquitin ligases are the crucial constituents by the advantage of recognizing and presenting proteins to proteasomes for proteolysis. As the major regulators of protein homeostasis, E3 ligases are indispensable to proper cell manners in diverse systems, and they are well described in physiological bone growth and bone metabolism. Pathologically, classic bone-related diseases such as metabolic bone diseases, arthritis, bone neoplasms and bone metastasis of the tumor, etc., were also depicted in a UPS-dependent manner. Therefore, skeletal system is versatilely regulated by UPS and it is worthy to summarize the underlying mechanism. Furthermore, based on the current status of treatment, normal or pathological osteogenesis and tumorigenesis elaborated in this review highlight the clinical significance of UPS research. As a strategy possibly remedies the limitations of UPS treatment, emerging PROTAC was described comprehensively to illustrate its potential in clinical application. Altogether, the purpose of this review aims to provide more evidence for exploiting novel therapeutic strategies based on UPS for bone associated diseases.

The Role of NEDD4-1 in Tumorigenesis of the Digestive System / 中国生物化学与分子生物学报

Neural precursor cell Expressed‚Developmentally Down-regulated protein 4 (NEDD4-1‚ also known as NEDD4 in some papers) is a tumor-related protein that has attracted much attention in recent years. It belongs to the E3 HECT (homologous to E6 associated protein C terminus) ubiquitin ligase‚ which could ubiquitinate various proteins that are subsequently degraded in lysosomes or proteasomes‚ or mediate their nuclear-cytoplasmic translocation‚ or indirectly affect various signaling pathways of different malignant tumors. With the deepening of a large number of tumor-related experiments‚ it has been found that NEDD4-1 can affect the biological behavior of tumors by regulating cell cycle‚ invasion and metastasis of cancer cells‚ antagonize drug resistance and many other pathways. In digestive system tumors‚ NEDD4-1 mainly promotes the proliferation‚ invasion and migration of hepatocellular carcinoma through multiple pathways such as PTEN/ PI3K/ Akt‚ TGF-β‚ Hippo and LDLRAD4. In pancreatic cancer‚ NEDD4-1 acted as an oncogene in the PI3K/ Akt signaling pathway‚ but acted as a tumor suppressor gene in the Myc-Sirt2 signaling circuit. In gastric and colorectal cancer‚ the NEDD4-1-related signaling pathways are different from other digestive system tumors. NEDD4-1 promotes gastric cancer progression and metastasis (via the EGFR signaling pathway) and inhibits colorectal cancer tumor growth (via the Wnt signaling pathway) independently of the PTEN/ PI3K/ Akt pathway. NEDD4-1 has become a hot research direction for therapeutic purposes. In this paper‚ we summarize the functions‚ signaling pathways and potential inhibitors of NEDD4-1 in different digestive system tumors‚ and discuss the relationship between NEDD4-1 and different signaling pathways‚ aiming to provide important reference data for cancer therapy.

Effect of Gandou Decoction on Mitophagy in Toxic Milk （TX） Mouse Model of Wilson Disease Based on Pink1/Parkin Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effects of Gandou decoction （GDD） on the mitophagy of hippocampal neurons in toxic milk （TX） mouse model of Wilson disease and explore the protective mechanism of GDD against neuron injury through the PTEN induced kinase 1 （Pink1） /E3 ubiquitin ligase （Parkin） pathway. MethodSixty mice were randomly divided into a blank group， a model group， a penicillamine group （0.09 g·kg-1）， and low- （5.5 g·kg-1）， medium- （11 g·kg-1）， and high-dose （22 g·kg-1） GDD groups， and treated correspondingly by gavage for 8 weeks. Morris water maze， traction test， and pole test were used for the evaluation of animal behaviors. Hematoxylin-eosin （HE） staining and transmission electron microscopy were used to observe cell apoptosis， ultrastructure， autophagy， and mitochondrial structure. The levels of superoxide dismutase （SOD）， reactive oxygen species （ROS）， and malondialdehyde （MDA） were detected by enzyme-linked immunosorbent assay （ELISA）. Real-time fluorescence-based quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of Pink1， Parkin， autophagy-associated protein Beclin-1， microtubule-associated protein 1 light chain 3Ⅱ （LC3Ⅱ）， and p62. Western blot was conducted to detect the protein expression of Pink1， Parkin， Beclin-1， LC3Ⅱ/Ⅰ， and p62. ResultCompared with the blank group， the model group showed prolonged escape latency， decreased times of platform crossing， lower score in the traction test， and longer pole climbing time （P<0.01）. Compared with the model group， the medium- and high-dose GDD groups and the penicillamine group showed shortened escape latencies， increased times of platform crossing， higher scores in the traction test， and shortened pole climbing time （P<0.01）. Compared with the blank group， the model group displayed severely damaged neurons and increased autophagosomes. Compared with the model group， the medium- and high-dose GDD groups and the penicillamine group showed improved neuron damage and reduced autophagosomes. The levels of ROS and MDA were higher and SOD was lower in the model group than those in the blank group （P<0.01）， while the levels of the above indicators were reversed by GDD intervention as compared with the model group （P<0.01）. Compared with the blank group， the model group exhibited up-regulated mRNA and protein expression of Pink1， Parkin， LC3Ⅱ， and Beclin-1 and down-regulated p62 （P<0.05）. Compared with the model group， the medium- and high-dose GDD groups showed reduced mRNA and protein expression of Pink1， Parkin， LC3Ⅱ， and Beclin-1 and increased p62 （P<0.05， P<0.01）. ConclusionGDD can significantly inhibit the excessive mitophagy in neurons of TX mice and protect neurons from damage. The mechanism may be related to the regulation of the Pink1/Parkin pathway.

mTOR-targeted cancer therapy: great target but disappointing clinical outcomes, why? / 医学前沿

The mammalian target of rapamycin (mTOR) critically regulates several essential biological functions, such as cell growth, metabolism, survival, and immune response by forming two important complexes, namely, mTOR complex 1 (mTORC1) and complex 2 (mTORC2). mTOR signaling is often dysregulated in cancers and has been considered an attractive cancer therapeutic target. Great efforts have been made to develop efficacious mTOR inhibitors, particularly mTOR kinase inhibitors, which suppress mTORC1 and mTORC2; however, major success has not been achieved. With the strong scientific rationale, the intriguing question is why cancers are insensitive or not responsive to mTOR-targeted cancer therapy in clinics. Beyond early findings on induced activation of PI3K/Akt, MEK/ERK, and Mnk/eIF4E survival signaling pathways that compromise the efficacy of rapalog-based cancer therapy, recent findings on the essential role of GSK3 in mediating cancer cell response to mTOR inhibitors and mTORC1 inhibition-induced upregulation of PD-L1 in cancer cells may provide some explanations. These new findings may also offer us the opportunity to rationally utilize mTOR inhibitors in cancer therapy. Further elucidation of the biology of complicated mTOR networks may bring us the hope to develop effective therapeutic strategies with mTOR inhibitors against cancer.

Parkin: an Extremely Versatile E3 Ubiquitin Ligase / 中国生物化学与分子生物学报

Parkin, also known as PARK2, has been closely related to Parkinson's disease (PD) since its discovery. It is considered to be a neuroprotective gene. With the in-depth understanding for its structure, Parkin has been unveiled as an E3 ubiquitin ligase. Parkin is involved in the regulation of cell cycle, mitochondrial homeostasis, energy metabolism and other cellular processes, and is closely related to many diseases. It even plays completely opposite roles in the same pathway, namely cell proliferation and apoptosis, indicating that this must be a gene with an extremely broad and important role. This article summarizes the discovery and structure of Parkin and its self-inhibiting characteristics, focusing on the ubiquitination process that it participates in as E3 ubiquitin ligase and the resulting autophagy, protein degradation, changes in protein subcellular localization and protein interaction. These may all serve as the basis for Parkin to prevent PD and suppress tumors. On this basis, two reasons for Parkin abnormalities leading to PD are summarized: abnormal protein quality control and mitochondrial dysfunction, and extended to cardiovascular and kidney diseases caused by the abnormality of Parkin due to mitochondrial dysfunction. The internal connection between Parkin and cancer is also introduced from the aspects of Parkin as a tumor suppressor, regulating cell cycle, apoptosis and metastasis, oxidative stress and energy metabolism. By maintaining the active state of Parkin or enhancing its expression, it may be possible to improve the condition of PD patients. But the mechanism of Parkin's inhibition of tumor growth remains to be deciphered, and the potential role of Parkin in mediating the relationship between PD and cancer risk should be strengthened. These follow-up in-depth studies and their role in the diagnosis and treatment of related diseases and application of target molecules laid the foundation.

Localization of Melanoma-associated Antigen MAGEA1 and Identification of Its Interacting Proteins / 中国生物化学与分子生物学报

Melanoma associated antigen family A1 (MAGEA1) is expressed in germ cells and tumors of various histological origins, but its mechanism is still unclear. In this study, the eukaryotic recombinant MAGEA1 expression plasmids with Flag or GFP tags were constructed and transfected into HeLa and HEK293T cells. Western blotting, immunocytochemistry, co-immunoprecipitation, nuclear protein and cytoplasmic protein separation, and mitochondrial isolation were used to detect the expression and location of MAGEA1 and its interaction with other proteins in cells. The results of immunocytochemistry (ICC) and Western blotting showed that the overexpressed MAGEA1 was mainly localized in the cytoplasm and partially co-localized with mitochondria. Co-immunoprecipitation experiments verified the interactions between MAGEA1 and TRIM31, SNW1, HDAC1, and found that MAGEA1 may mainly interact with HDAC1 in the cytoplasm. The studies above indicate that MAGEA1 may be involved in different cellular biological processes and co-localize with mitochondria. It interacts with TRIM31, SNW1 and HDAC1, while MAGEA1 may mainly interact with HDAC1 in the cytoplasm. We propose that it may be involved in protein ubiquitination and the Notch signaling pathway. The results of this study laid an experimental foundation for the subsequent in-depth study of the mechanism of MAGEA1.

Emerging Roles of the TRIM Family in Autophagy / 中国生物化学与分子生物学报

Autophagy is a common cellular metabolic process, which is characterized by the formation of double membrane structures named autophagosomes to degrade intracellular components or invading foreign substances to maintain cellular homeostasis. Autophagy is crucial for maintaining cell homeostasis. The dysfunction of autophagy is closely related to the occurrence and development of various diseases, including tumors, neurodegenerative diseases, viral infection, immune diseases and so on. Autophagy may be a potential therapeutic target for these diseases. Therefore, the investigation of autophagy regulation is a hot issue in life science and medical research. The TRIM (tripartite motif-containing proteins) family is a set of proteins with E3 ubiquitin ligase activity and usually contains three conserved domains, a RING zinc finger structure, a B-box structure and a coiled helix domain. Many TRIM family members have been found to play important roles in autophagy regulation, the mechanism of which include modulating autophagy-related signaling pathways, regulating autophagy core molecules and acting as autophagy receptors, etc. TRIMs participate in many biological pathways through regulating autophagy, such as immunity, virus infection and tumors. This review covers the role of TRIM proteins in regulating autophagy, the molecular mechanism and the corresponding biological effects.

Progress of drugs targeting E3 ubiquitin ligase / 中国药理学通报

The ubiquitin-proteasome pathway is one of the most important pathways of cell protein degradation in eukaryotes, and plays an important role in the regulation of cell proliferation, differentiation, apoptosis, DNA repair and other physiological activities. E3 ubiquitin ligase is the major component of ubiquitinproteasome system, which is responsible for substrate recognition. The abnormal regulation of E3 ubiquitin ligase may cause many diseases such as cancer, Alzheimer's disease and Parkinson's disease. Here, we summarizes the progress of drugs targeting E3 ubiquitin ligase in cancer, Alzheimer's disease, Parkinson's disease, diabetic complications, atherosclerosis, and inflammatory bowel diseases. At present, only a few of small molecule antagonists or agonists targeting E3 ubiquitin ligase are under development. The study of natural products in China is leading the way in the world, and numerous natural products have been identified for pharmacological effects on E3 ubiquitin ligase, which may open up a new avenue for multiple complex diseases.

Mechanism of Effect of Ubiquitination Modification Mediated by E3 Ubiquitin Ligase in Inflammatory Bowel Disease / 胃肠病学

Ubiquitination is the important type of post-translational modification of proteins, and its main effects include protein degradation and functional regulation. Studies have shown that abnormal ubiquitination is involved in the development and progress of inflammatory bowel disease (IBD), which makes some ubiquitinases and their antibodies having the potential to be important markers for clinical diagnosis and evaluation of severity of Crohn's disease (CD). This article reviewed the research on mechanism of effect of ubiquitination modification mediated by E3 ubiquitin ligase in IBD.

Degradation of proteins by PROTACs and other strategies

Blocking the biological functions of scaffold proteins and aggregated proteins is a challenging goal. PROTAC proteolysis-targeting chimaera (PROTAC) technology may be the solution, considering its ability to selectively degrade target proteins. Recent progress in the PROTAC strategy include identification of the structure of the first ternary eutectic complex, extra-terminal domain-4-PROTAC-Von-Hippel-Lindau (BRD4-PROTAC-VHL), and PROTAC ARV-110 has entered clinical trials for the treatment of prostate cancer in 2019. These discoveries strongly proved the value of the PROTAC strategy. In this perspective, we summarized recent meaningful research of PROTAC, including the types of degradation proteins, preliminary biological data in vitro and in vivo, and new E3 ubiquitin ligases. Importantly, the molecular design, optimization strategy and clinical application of candidate molecules are highlighted in detail. Future perspectives for development of advanced PROTAC in medical fields have also been discussed systematically.

Role of RNF87 in predicting the prognosis of human hepatocellular carcinoma patients / 中国现代普通外科进展

Objective:To investigate the level and prognostic significance of RNF87 in human hepatocellular carcinoma.Methods:Detected the expression of RNF87 in 98 HCC tissues by immunohistochemistry and Western Blot.According to the clinical data of the patients,we analyzed the relationship between RNF87 level and the prognosis of the HCC patients.Results:The level of RNF87 in HCC tissues is down-regulated,compared with the adjacent tissues.And the expression of RNF87 was significantly related to the prognosis of HCC patients.Besides,the lower level of RNF87 was also obviously related with microvascular invasion.Conclusions:The down-regulated level of RNF87 may be one of the risk factors of human hepatocellular carcinoma progression;RNF87 maybe one of potential tumor suppressors;the level of RNF87 can be used as an indicator to predict the prognosis of HCC patients.

Progress in research on the novel cancer therapeutic target Cdc20 / 药学学报

The anaphase promoting complex (APC) regulates cell cycle progression by forming two functionally distinct E3 ubiquitin ligase complexes, APCCdc20 activated by cell division cycle protein 20 (Cdc20) and APCCdh1 activated by Cdc20 homologue 1 (Cdh1), respectively. Cdc20 and Cdh1 have different functions in the occurrence and development of the tumor. Cdc20 is a cancer promoter while Cdh1 suppresses tumorigenesis. Emerging evidence has begun to reveal that Cdc20 has positive functions in tumorigenesis, the overexpression of Cdc20 has been observed in many cancers. Currently, Cdc20 inhibitors, mostly non-specific inhibitors except apcin, not only block the combination between Cdc20 and APC, also block the combination between Cdh1 and APC, which leads to a poor selectivity. In this paper, the Cdc20 role in the development and process of cancers and its inhibitors are reviewed.

Loss of Ovarian Function Results in Increased Loss of Skeletal Muscle in Arthritic Rats / Perda da função ovariana resulta em perda de músculo esquelético aumentada em ratos artríticos

Objective We studied the effects of loss of ovarian function (ovariectomy) onmuscle mass of gastrocnemius and themRNA levels of IGF-1, atrogin-1, MuRF-1, andmyostatin in an experimental model of rheumatoid arthritis in rats. Methods We randomly allocated 24 female Wistar rats (9 weeks, 195.3±17.4 grams) into four groups: control (CT-Sham; n = 6); rheumatoid arthritis (RA; n = 6); ovariectomy without rheumatoid arthritis (OV; n = 6); ovariectomy with rheumatoid arthritis (RAOV; n = 6). We performed the ovariectomy (OV and RAOV) or Sham (CTSham or RA) procedures at the same time, fifteen days before the rheumatoid arthritis induction. The RA and RAOV groups were immunized and then were injected with Met- BSA in the tibiotarsal joint. After 15 days of intra-articular injections the animals were euthanized. We evaluated the external manifestations of rheumatoid arthritis (perimeter joint) as well as animal weight, and food intake throughout the study. We also analyzed the cross-sectional areas (CSA) of gastrocnemius muscle fibers in 200 fibers (H&E method). In the gastrocnemius muscle, we analyzed mRNA expression by quantitative real time PCR followed by the Livak method (ΔΔCT). Results The rheumatoid arthritis induced reduction in CSA of gastrocnemius muscle fibers. The RAOV group showed a lower CSA of gastrocnemius muscle fibers compared to RA and CT-Sham groups. Skeletal muscle IGF-1 mRNA increased in arthritics and ovariectomized rats. The increased IGF-1 mRNA was higher in OV groups than in the RA and RAOV groups. Antrogin-1 mRNA also increased in the gastrocnemius muscle of arthritic and ovariectomized rats. However, the increased atrogin-1 mRNA was higher in RAOV groups than in the RA and OV groups. Gastrocnemius muscle MuRF-1 mRNA increased in the OVand RAOVgroups, but not in the RA and Shamgroups. However, the RAOV group showed higher MuRF-1 mRNA than the OV group. The myostatin gene expression was similar in all groups. Conclusion Loss of ovarian function results in increased loss of skeletal musclerelated ubiquitin ligases atrogin-1 and MuRF-1 in arthritic rats.

Objetivo Foram estudados os efeitos da perda da função ovariana (ovariectomia) sobre músculo esquelético e os níveis de RNAm de IGF-1, atrogina-1, MuRF-1, e de miostatina em modelo experimental de artrite reumatóide em ratos. Métodos 24 ratos Wistar (9 semanas, 195,3±17,4 gramas) foram distribuídos aleatoriamente em quatro grupos: controle (CT-Sham, n = 6); artrite reumatóide (RA, n = 6); ovariectomia sem artrite reumatóide (OV; n = 6); ovariectomia com artrite reumatóide (RAOV; n = 6). Os procedimentos da ovariectomia (OV e RAOV) ou simulação da ovariectomia (CT-Shamou RA) foramrealizados aomesmo tempo, quinze dias antes da indução da artrite reumatóide. Os grupos RA e RAOV foramimunizados e, em seguida, foram injetados com Met-BSA na articulação tibiotársica. Após 15 dias das injeções intra-articulares, os animais foram eutanasiados. Foram avaliadas as manifestações externas da artrite reumatóide (perimetria articular), bem como o peso dos animais e a ingestão de alimentos ao longo do estudo. Além disso, as áreas de secção transversa (CSA) do músculo gastrocnêmio foram analisadas em 200 fibras (método H & E). No músculo gastrocnêmio, a expressão de RNAm foi analisada por PCR quantitativo em tempo real, seguido pelo método Livak (ΔΔCT). Resultados A artrite reumatoide reduziu a CSA das fibras do músculo gastrocnêmio. O grupo RAOV mostrou uma CSA menor nas fibras do músculo gastrocnêmio em comparação com os grupos RA e CT-Sham. O RNAm do IGF-1 do músculo esquelético aumentou nos ratos artríticos e ovariectomizados. O RNAm do IGF-1 foi maior nos grupos OV do que nos grupos RA e RAOV. A expressão de antrogina-1 também aumentou no músculo gastrocnêmio dos ratos artríticos e ovariectomizados. No entanto, o aumento do RNAm da atrogina-1 foi maior no grupo RAOV do que nos grupos RA e OV. O RNAm da MuRF-1 aumentou nos grupos OV e RAOV, mas não nos grupos RA e CT-Sham. Porém, o grupo RAOV apresentou maior expressão gênica de MuRF-1 do que o grupo OV. A expressão do gene da miostatina foi semelhante em todos os grupos. Conclusão A perda de função ovariana resulta em perda de músculo esquelético associado às ubiquitina-ligases atrogina-1 e MuRF-1 em ratos artríticos.

The Role of E3 Ubiquitin Ligase Smurfs in BMP and TGF-bata Signaling in Bone Cells / 生物化学与生物物理进展

The ubiquitin-proteasome system is composed by multiple enzymes which are ubiquitously expressed in mammalian cells and plays an essential role in a variety of biological processes. As key members of the protein degradation enzymatic system, the function of E3 ubiquitin ligases has been extensively investigated. BMP and TGF-? are critical molecules that regulate proliferation, differentiation and apoptosis of osteoblasts and chondrocytes through different signaling pathways. Resent findings indicate that the ubiquitin-proteasome system functions as a key regulator in bone cells and the E3 ligase mediates the proteolytic degradation of critical molecules in BMP and TGF-? signaling pathways. Recent progress on studies of HECT domain E3 ligase, Smurf, in osteoblasts and chondrocytes were summarized. The regulatory role of Smurf1 and Smurf2 in BMP and TGF-? signaling and osteoblast and chondrocyte function has been reviewed.