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The NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome is a crucial component of the innate immune system that initiates inflammatory responses. Posttranslational modifications (PTMs) of NLRP3, including ubiquitination and phosphorylation, control inflammasome activation and determine the intensity of inflammation. However, the role of other PTMs in controlling NLRP3 inflammasome activation remains unclear. This study found that TLR priming induced NLRP3 ISGylation (a type of PTM in which ISG15 covalently binds to the target protein) to stabilize the NLRP3 protein. Viral infection, represented by SARS-COV-2 infection, and type I IFNs induced expression of ISG15 and the predominant E3 ISGylation ligases HECT domain- and RCC1-like domain-containing proteins (HERCs; HERC5 in humans and HERC6 in mice). HERCs promoted NLRP3 ISGylation and inhibited K48-linked ubiquitination and proteasomal degradation, resulting in the enhancement of NLRP3 inflammasome activation. Concordantly, Herc6 deficiency ameliorated NLRP3-dependent inflammation as well as hyperinflammation caused by viral infection. The results illustrate the mechanism by which type I IFNs responses control inflammasome activation and viral infection-induced aberrant NLRP3 activation. This work identifies ISGylation as a PTM of NLRP3, revealing a priming target that modulates NLRP3-dependent immunopathology., Introduction The NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome is a vital component of the innate immune system that senses both pathogen-associated molecular patterns (PAMPs) from microbes and [...]

Hormone replacement therapy (HRT) is not recommended for treating learning and memory decline in menopausal women because it exerts adverse effects by activating classic estrogen receptors ERa and ERp. The membrane estrogen receptor G protein-coupled receptor 30 (GPR30) has been reported to be involved in memory modulation; however, the underlying mechanisms are poorly understood. Here, we found that GPR30 deletion in astrocytes, but not in neurons, impaired learning and memory in female mice. Astrocytic GPR30 depletion induced A1 phenotype transition, impairing neuronal function. Further exploration revealed that Praja1 (PJA1), a RING ubiquitin ligase, mediated the effects of astrocytic GPR30 on learning and memory by binding to Serpina3n, which is a molecular marker of neuroinflammation in astrocytes. GPR30 positively modulated PJA1 expression through the CREB signaling pathway in cultured murine and human astrocytes. Additionally, the mRNA levels of GPR30 and PJA1 were reduced in exosomes isolated from postmenopausal women while Serpina3n levels were increased in the plasma. Together, our findings suggest a key role for astrocytic GPR30 in the learning and memory abilities of female mice and identify GPR30/PJA1/Serpina3n as potential therapeutic targets for learning and memory loss in peri- and postmenopausal women., Introduction The process of learning and memory is how people acquire, encode, store, retain and later retrieve information in the brain. For perimenopausal women, learning and memory abilities decline rapidly [...]

The ubiquitin-proteasome system (UPS) comprises a network of enzymes that is responsible for maintaining cellular protein homeostasis. The therapeutic potential of this pathway has been validated by the clinical successes of a number of UPS modulators, including proteasome inhibitors and immunomodulatory imide drugs (IMiDs). Here we identified TAK-243 (formerly known as MLN7243) as a potent, mechanism-based small-molecule inhibitor of the ubiquitin activating enzyme (UAE), the primary mammalian E1 enzyme that regulates the ubiquitin conjugation cascade. TAK-243 treatment caused depletion of cellular ubiquitin conjugates, resulting in disruption of signaling events, induction of proteotoxic stress, and impairment of cell cycle progression and DNA damage repair pathways. TAK-243 treatment caused death of cancer cells and, in primary human xenograft studies, demonstrated antitumor activity at tolerated doses. Due to its specificity and potency, TAK-243 allows for interrogation of ubiquitin biology and for assessment of UAE inhibition as a new approach for cancer treatment., Author(s): Marc L Hyer [1]; Michael A Milhollen (corresponding author) [1]; Jeff Ciavarri [1]; Paul Fleming [1]; Tary Traore [1]; Darshan Sappal [1]; Jessica Huck [1]; Judy Shi [1]; James [...]

Autosomal-recessive juvenile Parkinsonism (AR-JP) is caused by mutations in a number of PARK genes, in particular the genes encoding the E3 ubiquitin ligase Parkin (PARK2, also known as PRKN) and its upstream protein kinase PINK1 (also known as PARK6). PINK1 phosphorylates both ubiquitin and the ubiquitin-like domain of Parkin on structurally protected Ser65 residues, triggering mitophagy. Here we report a crystal structure of a nanobody-stabilized complex containing Pediculus humanus corporis (Ph)PINK1 bound to ubiquitin in the C-terminally retracted (Ub-CR) conformation. The structure reveals many peculiarities of PINK1, including the architecture of the C-terminal region, and reveals how the N lobe of PINK1 binds ubiquitin via a unique insertion. The flexible Ser65 loop in the Ub-CR conformation contacts the activation segment, facilitating placement of Ser65 in a phosphate-accepting position. The structure also explains how autophosphorylation in the N lobe stabilizes structurally and functionally important insertions, and reveals the molecular basis of AR-JP-causing mutations, some of which disrupt ubiquitin binding., Author(s): Alexander F. Schubert [1]; Christina Gladkova [1]; Els Pardon [2, 3]; Jane L. Wagstaff [1]; Stefan M. V. Freund [1]; Jan Steyaert [2, 3]; Sarah L. Maslen [1]; David [...]

Non-alcoholic steatohepatitis (NASH) is an increasingly prevalent liver pathology that can progress from non-alcoholic fatty liver disease (NAFLD), and it is a leading cause of cirrhosis and hepatocellular carcinoma. There is currently no pharmacological therapy for NASH. Defective lysosome-mediated protein degradation is a key process that underlies steatohepatitis and a well-recognized drug target in a variety of diseases; however, whether it can serve as a therapeutic target for NAFLD and NASH remains unknown. Here we report that transmembrane BAX inhibitor motif-containing 1 (TMBIM1) is an effective suppressor of steatohepatitis and a previously unknown regulator of the multivesicular body (MVB)-lysosomal pathway. Tmbim1 expression in hepatocytes substantially inhibited high-fat diet-induced insulin resistance, hepatic steatosis and inflammation in mice. Mechanistically, Tmbim1 promoted the lysosomal degradation of toll-like receptor 4 by cooperating with the ESCRT endosomal sorting complex to facilitate MVB formation, and the ubiquitination of Tmbim1 by the E3 ubiquitin ligase Nedd4l was required for this process. We also found that overexpression of Tmbim1 in the liver effectively inhibited a severe form of NAFLD in mice and NASH progression in monkeys. Taken together, these findings could lead to the development of promising strategies to treat NASH by targeting MVB regulators to properly orchestrate the lysosome-mediated protein degradation of key mediators of the disease., Author(s): Guang-Nian Zhao [1, 2, 3, 4, 5]; Peng Zhang [1, 2, 3, 4]; Jun Gong [2, 3, 4, 5]; Xiao-Jing Zhang [2, 3, 4, 6]; Pi-Xiao Wang [2, 3, [...]

ABSTRACT Background and Purpose: Aberrant expression of β-catenin is highly associated with progression of various cancers including head and neck cancer (HNC). Green tea is most commonly used beverage in [...]

Thalidomide and its derivatives, lenalidomide and pomalidomide, are immune modulatory drugs (IMiDs) used in the treatment of haematologic malignancies (1,2). IMiDs bind CRBN, the substrate receptor of the CUL4-RBX1-DDB1-CRBN (also [...]