The Giant HECT E3 Ubiquitin Ligase HERC1 Is Aberrantly Expressed in Myeloid Related Disorders and It Is a Novel BCR-ABL1 Binding Partner

HERC E3 subfamily members are parts of the E3 ubiquitin ligases and key players for a wide range of cellular functions. Though the involvement of the Ubiquitin Proteasome System in blood disorders has been broadly studied, so far the role of large HERCs in this context remains unexplored. In the present study we examined the expression of the large HECT E3 Ubiquitin Ligase, HERC1, in blood disorders. Our findings revealed that HERC1 gene expression was severely downregulated both in acute and in chronic myelogenous leukemia at diagnosis, while it is restored after complete remission achievement. Instead, in Philadelphia the negative myeloproliferative neoplasm HERC1 level was peculiarly controlled, being very low in Primary Myelofibrosis and significantly upregulated in those Essential Thrombocytemia specimens harboring the mutation in the calreticulin gene. Remarkably, in CML cells HERC1 mRNA level was associated with the BCR-ABL1 kinase activity and the HERC1 protein physically interacted with BCR-ABL1. Furthermore, we found that HERC1 was directly tyrosine phosphorylated by the ABL kinase. Overall and for the first time, we provide original evidence on the potential tumor-suppressing or -promoting properties, depending on the context, of HERC1 in myeloid related blood disorders.

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The Molecular and Pathophysiological Functions of Members of the LNX/PDZRN E3 Ubiquitin Ligase Family

Molecules ◽

10.3390/molecules25245938 ◽

2020 ◽

Vol 25 (24) ◽

pp. 5938

Author(s):

Jeongkwan Hong ◽

Minho Won ◽

Hyunju Ro

Keyword(s):

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Ligases ◽

Ubiquitin Proteasome System ◽

E3 Ubiquitin Ligases ◽

Intercellular Signaling ◽

Cellular Functions ◽

Ring Type ◽

Ubiquitin Ligase Activity ◽

Ubiquitin Proteasome

The ligand of Numb protein-X (LNX) family, also known as the PDZRN family, is composed of four discrete RING-type E3 ubiquitin ligases (LNX1, LNX2, LNX3, and LNX4), and LNX5 which may not act as an E3 ubiquitin ligase owing to the lack of the RING domain. As the name implies, LNX1 and LNX2 were initially studied for exerting E3 ubiquitin ligase activity on their substrate Numb protein, whose stability was negatively regulated by LNX1 and LNX2 via the ubiquitin-proteasome pathway. LNX proteins may have versatile molecular, cellular, and developmental functions, considering the fact that besides these proteins, none of the E3 ubiquitin ligases have multiple PDZ (PSD95, DLGA, ZO-1) domains, which are regarded as important protein-interacting modules. Thus far, various proteins have been isolated as LNX-interacting proteins. Evidence from studies performed over the last two decades have suggested that members of the LNX family play various pathophysiological roles primarily by modulating the function of substrate proteins involved in several different intracellular or intercellular signaling cascades. As the binding partners of RING-type E3s, a large number of substrates of LNX proteins undergo degradation through ubiquitin-proteasome system (UPS) dependent or lysosomal pathways, potentially altering key signaling pathways. In this review, we highlight recent and relevant findings on the molecular and cellular functions of the members of the LNX family and discuss the role of the erroneous regulation of these proteins in disease progression.

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Ubiquitin-dependent regulation of immune and inflammatory signaling pathways

Acta Crystallographica Section A Foundations and Advances ◽

10.1107/s2053273314097587 ◽

2014 ◽

Vol 70 (a1) ◽

pp. C241-C241

Author(s):

Katrin Rittinger

Keyword(s):

Signaling Pathways ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

High Specificity ◽

Cellular Functions ◽

Inflammatory Signaling ◽

Ubiquitin Chain ◽

E3 Ligases ◽

Wide Range ◽

Chain Synthesis

Modification of proteins with ubiquitin is a key mechanism for the regulation of a wide range of cellular functions. The outcome of the modification is determined by the way ubiquitin molecules are linked to each other. Linear (M1-linked) ubiquitin chains play an important role in the regulation of immune and inflammatory signaling pathways and contribute to the activation of NF-κB. They are synthesized by the E3 ubiquitin ligase LUBAC (linear ubiquitin chain assembly complex) that is composed of at least three subunits named HOIL-1L, HOIP and SHARPIN. LUBAC belongs to the RBR (RING-inbetween-RING) family of E3 ligases that combine the properties of RING and HECT ligases and act as RING/HECT hybrids. Indeed, we have recently shown that linear ubiquitin chain synthesis proceeds via ubiquitin thioester intermediate formed by the HOIP subunit before subsequent transfer onto the target. I will present a combination of structural and biochemical data that provide a molecular explanation how this unusual E3 ligase complex promotes the synthesis of linear ubiquitin chains with high specificity, regardless of the E2 conjugating enzyme it works with.

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Proteomic analysis identifies the E3 ubiquitin ligase Pdzrn3 as a regulatory target of Wnt5a-Ror signaling

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2104944118 ◽

2021 ◽

Vol 118 (25) ◽

pp. e2104944118

Author(s):

Sara E. Konopelski Snavely ◽

Michael W. Susman ◽

Ryan C. Kunz ◽

Jia Tan ◽

Srisathya Srinivasan ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Large Scale ◽

Glycogen Synthase ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Proteasome System ◽

Dependent Manner ◽

Casein Kinase 1 ◽

Downstream Signaling ◽

Proteomic Screen ◽

Responsive Element

Wnt5a-Ror signaling is a conserved pathway that regulates morphogenetic processes during vertebrate development [R. T. Moon et al., Development 119, 97–111 (1993); I. Oishi et al., Genes Cells 8, 645–654 (2003)], but its downstream signaling events remain poorly understood. Through a large-scale proteomic screen in mouse embryonic fibroblasts, we identified the E3 ubiquitin ligase Pdzrn3 as a regulatory target of the Wnt5a-Ror pathway. Upon pathway activation, Pdzrn3 is degraded in a β-catenin–independent, ubiquitin-proteasome system–dependent manner. We developed a flow cytometry-based reporter to monitor Pdzrn3 abundance and delineated a signaling cascade involving Frizzled, Dishevelled, Casein kinase 1, and Glycogen synthase kinase 3 that regulates Pdzrn3 stability. Epistatically, Pdzrn3 is regulated independently of Kif26b, another Wnt5a-Ror effector. Wnt5a-dependent degradation of Pdzrn3 requires phosphorylation of three conserved amino acids within its C-terminal LNX3H domain [M. Flynn, O. Saha, P. Young, BMC Evol. Biol. 11, 235 (2011)], which acts as a bona fide Wnt5a-responsive element. Importantly, this phospho-dependent degradation is essential for Wnt5a-Ror modulation of cell migration. Collectively, this work establishes a Wnt5a-Ror cell morphogenetic cascade involving Pdzrn3 phosphorylation and degradation.

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Impaired muscle morphology in a Drosophila model of myosin storage myopathy was supressed by overexpression of an E3 ubiquitin ligase

Disease Models & Mechanisms ◽

10.1242/dmm.047886 ◽

2020 ◽

Vol 13 (12) ◽

pp. dmm047886

Author(s):

Martin Dahl-Halvarsson ◽

Montse Olive ◽

Malgorzata Pokrzywa ◽

Michaela Norum ◽

Katarina Ejeskär ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Mutant Allele ◽

Body Movement ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Proteasome System ◽

Muscle Disease ◽

Muscle Morphology ◽

Severe Impairment ◽

Myosin Storage Myopathy ◽

The Impact

ABSTRACTMyosin is vital for body movement and heart contractility. Mutations in MYH7, encoding slow/β-cardiac myosin heavy chain, are an important cause of hypertrophic and dilated cardiomyopathy, as well as skeletal muscle disease. A dominant missense mutation (R1845W) in MYH7 has been reported in several unrelated cases of myosin storage myopathy. We have developed a Drosophila model for a myosin storage myopathy in order to investigate the dose-dependent mechanisms underlying the pathological roles of the R1845W mutation. This study shows that a higher expression level of the mutated allele is concomitant with severe impairment of muscle function and progressively disrupted muscle morphology. The impaired muscle morphology associated with the mutant allele was suppressed by expression of Thin (herein referred to as Abba), an E3 ubiquitin ligase. This Drosophila model recapitulates pathological features seen in myopathy patients with the R1845W mutation and severe ultrastructural abnormalities, including extensive loss of thick filaments with selective A-band loss, and preservation of I-band and Z-disks were observed in indirect flight muscles of flies with exclusive expression of mutant myosin. Furthermore, the impaired muscle morphology associated with the mutant allele was suppressed by expression of Abba. These findings suggest that modification of the ubiquitin proteasome system may be beneficial in myosin storage myopathy by reducing the impact of MYH7 mutation in patients.

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p97 Negatively Regulates NRF2 by Extracting Ubiquitylated NRF2 from the KEAP1-CUL3 E3 Complex

Molecular and Cellular Biology ◽

10.1128/mcb.00660-16 ◽

2017 ◽

Vol 37 (8) ◽

Cited By ~ 36

Author(s):

Shasha Tao ◽

Pengfei Liu ◽

Gang Luo ◽

Montserrat Rojo de la Vega ◽

Heping Chen ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Protein Complexes ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Proteasome System ◽

Proteasomal Degradation ◽

Ubiquitin Ligase Complex ◽

Ubiquitin Proteasome ◽

Client Proteins

ABSTRACT Activation of the stress-responsive transcription factor NRF2 is the major line of defense to combat oxidative or electrophilic insults. Under basal conditions, NRF2 is continuously ubiquitylated by the KEAP1-CUL3-RBX1 E3 ubiquitin ligase complex and is targeted to the proteasome for degradation (the canonical mechanism). However, the path from the CUL3 complex to ultimate proteasomal degradation was previously unknown. p97 is a ubiquitin-targeted ATP-dependent segregase that extracts ubiquitylated client proteins from membranes, protein complexes, or chromatin and has an essential role in autophagy and the ubiquitin proteasome system (UPS). In this study, we show that p97 negatively regulates NRF2 through the canonical pathway by extracting ubiquitylated NRF2 from the KEAP1-CUL3 E3 complex, with the aid of the heterodimeric cofactor UFD1/NPL4 and the UBA-UBX-containing protein UBXN7, for efficient proteasomal degradation. Given the role of NRF2 in chemoresistance and the surging interest in p97 inhibitors to treat cancers, our results indicate that dual p97/NRF2 inhibitors may offer a more potent and long-term avenue of p97-targeted treatment.

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The E3 ubiquitin ligase UBR5 regulates centriolar satellite stability and primary cilia

Molecular Biology of the Cell ◽

10.1091/mbc.e17-04-0248 ◽

2018 ◽

Vol 29 (13) ◽

pp. 1542-1554 ◽

Cited By ~ 12

Author(s):

Robert F. Shearer ◽

Kari-Anne Myrum Frikstad ◽

Jessie McKenna ◽

Rachael A. McCloy ◽

Niantao Deng ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Developmental Disorders ◽

Primary Cilia ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Proteasome System ◽

Interacting Protein ◽

Robust Model ◽

Cilia Formation ◽

Cytoplasmic Organization ◽

Ubiquitin Proteasome

Primary cilia are crucial for signal transduction in a variety of pathways, including hedgehog and Wnt. Disruption of primary cilia formation (ciliogenesis) is linked to numerous developmental disorders (known as ciliopathies) and diseases, including cancer. The ubiquitin–proteasome system (UPS) component UBR5 was previously identified as a putative positive regulator of ciliogenesis in a functional genomics screen. UBR5 is an E3 ubiquitin ligase that is frequently deregulated in tumors, but its biological role in cancer is largely uncharacterized, partly due to a lack of understanding of interacting proteins and pathways. We validated the effect of UBR5 depletion on primary cilia formation using a robust model of ciliogenesis, and identified CSPP1, a centrosomal and ciliary protein required for cilia formation, as a UBR5-interacting protein. We show that UBR5 ubiquitylates CSPP1, and that UBR5 is required for cytoplasmic organization of CSPP1-comprising centriolar satellites in centrosomal periphery, suggesting that UBR5-mediated ubiquitylation of CSPP1 or associated centriolar satellite constituents is one underlying requirement for cilia expression. Hence, we have established a key role for UBR5 in ciliogenesis that may have important implications in understanding cancer pathophysiology.

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Strategies for the identification of ubiquitin ligase inhibitors

Biochemical Society Transactions ◽

10.1042/bst0380132 ◽

2010 ◽

Vol 38 (1) ◽

pp. 132-136 ◽

Cited By ~ 25

Author(s):

Seth J. Goldenberg ◽

Jeffrey G. Marblestone ◽

Michael R. Mattern ◽

Benjamin Nicholson

Keyword(s):

Ubiquitin Ligase ◽

Therapeutic Target ◽

Therapeutic Strategy ◽

Ubiquitin Proteasome System ◽

Advantages And Disadvantages ◽

Wide Range ◽

Attractive Therapeutic Target ◽

Ubiquitin Proteasome ◽

Effective Therapeutic Strategy ◽

Protein Ligases

Dysregulation of the UPS (ubiquitin–proteasome system) has been implicated in a wide range of pathologies including cancer, neurodegeneration and viral infection. Inhibiting the proteasome has been shown to be an effective therapeutic strategy in humans; however, toxicity with this target remains high. E3s (Ub–protein ligases) represent an alternative attractive therapeutic target in the UPS. In this paper, we will discuss current platforms that report on E3 ligase activity and can detect E3 inhibitors, and underline the advantages and disadvantages of each approach.

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Antagonistic Regulation of Circadian Output and Synaptic Development by the E3 Ubiquitin Ligase JETLAG and the DYSCHRONIC-SLOWPOKE Complex

10.1101/769505 ◽

2019 ◽

Author(s):

Angelique Lamaze ◽

James E.C Jepson ◽

Oghenerukevwe Akpoghiran ◽

Kyunghee Koh

Keyword(s):

Neuromuscular Junction ◽

Ubiquitin Ligase ◽

Light Exposure ◽

E3 Ubiquitin Ligase ◽

List Type ◽

Cellular Functions ◽

Synaptic Development ◽

Channel Expression ◽

Pdz Protein ◽

Larval Neuromuscular Junction

SummaryCircadian output genes act downstream of the clock to promote rhythmic changes in behavior and physiology, yet their molecular and cellular functions are not well understood. Here we characterize an interaction between regulators of circadian entrainment, output and synaptic development in Drosophila that influences clock-driven anticipatory increases in morning and evening activity. We previously showed the JETLAG (JET) E3 Ubiquitin ligase resets the clock upon light exposure, while the PDZ protein DYSCHRONIC (DYSC) regulates circadian locomotor output and synaptic development. Surprisingly, we find that JET and DYSC antagonistically regulate synaptic development at the larval neuromuscular junction, and reduced JET activity rescues arrhythmicity of dysc mutants. Consistent with our prior finding that DYSC regulates SLOWPOKE (SLO) potassium channel expression, jet mutations also rescue circadian and synaptic phenotypes in slo mutants. Collectively, our data suggest that JET, DYSC and SLO promote circadian output in part by regulating synaptic morphology.HighlightsLoss of DYSC differentially impacts morning and evening oscillatorsReduced JET activity rescues the dysc and slo arrhythmic phenotypeReduced JET activity causes synaptic defects at the larval NMJJET opposes DYSC and SLO function at the NMJ synapse

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Investigating Patient Characteristics and Outcomes of Myelofibrosis Patients with Hyperleukocytosis

Blood ◽

10.1182/blood-2018-99-117834 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 3041-3041

Author(s):

Savan Shah ◽

Chetasi Talati ◽

Najla Al Ali ◽

Eric Padron ◽

David A Sallman ◽

...

Keyword(s):

Overall Survival ◽

Research Funding ◽

Management Strategies ◽

Myeloproliferative Neoplasm ◽

Primary Myelofibrosis ◽

Disease Diagnosis ◽

Extramedullary Hematopoiesis ◽

Myelogenous Leukemia ◽

Similar Proportion ◽

Increased Risk

Abstract Background: Primary myelofibrosis (PMF) is a myeloproliferative neoplasm that is marked by bone marrow fibrosis, cytopenias, extramedullary hematopoiesis, and a increased risk of acute leukemia. Leukocytosis is frequently seen and a white blood cell count (WBC) > 25,000/uL has been shown to convey an inferior prognosis. Rarely, patients develop severe leukocytosis (defined as WBC > 50,000/uL) either at presentation or during the course of their disease. Genomic characterization, management, and outcomes of these patients are not well defined. In this study, we aimed to further characterize PMF patients with severe leukocytosis and assess their clinical outcomes. Methods: We retrospectively reviewed PMF patients who presented to our institution between 2001 and 2018. We included patients who developed a WBC count > 50K at any time during their clinical course. Patients who developed acute myelogenous leukemia (AML) were censored at the time of disease diagnosis. Overall survival (OS) defined from first date that WBC documented as > 50K. Results: Among 493 PMF patients treated at our institution, 71 (14.4%) developed severe leukocytosis during the course of their disease. Ten (14%) had severe leukocytosis at the time of diagnosis and 30 (42%) developed it within 1 year of diagnosis and 40 (56%) developed it more than 1 year after diagnosis. Eight (11.3%) patients demonstrated peripheral blast percentage > 10% blasts at the time severe leukocytosis first documented. Compared to patients who did not develop severe leukocytosis, those with severe leukocytosis had an increased frequency of EZH2 (p < 0.001), RAS (p < 0.001), and KIT (p = 0.04) mutations. ASXL1 mutations were seen in a similar proportion of patients (p = 0.41). A similar proportion of patients were high risk by GIPSS (26% v 24%) prognostic model. From the time development of severe leukocytosis, the median overall survival (mOS) was 13.3 months. Median OS from diagnosis was significantly shorter for patients who developed severe leukocytosis (35.4 mo v 63.5 mo; p = 0.02) compared to those that did not. Eleven (15%) patients with severe leukocytosis developed AML compared to 38 (9%) patients who did not develop severe leukocytosis (p = 0.13). Median time to development of AML was 13.3 months from time of severe leukocytosis. At development of severe leukocytosis, 12 patients (17%) received hydrea (mOS 14.0 months) and 9 patients (13%) received ruxolitinib (mOS 21.2 months). The remaining cohort which either received no documented treatment, an alternative treatment or unknown treatment had a mOS of 5.3 months. This was not statistically significant. Ten patients (14%) ultimately underwent allogeneic stem cell transplantation after the development of hyperleukocytosis. Conclusion: Severe leukocytosis occurs rarely in primary myelofibrosis. RAS, KIT, and EZH2 mutations are enriched in these patients. Management strategies in these patients is varied and outcomes are poor. Further studies assessing the benefit of cytoreductive therapies in this population should be performed. Figure Figure. Disclosures Sallman: Celgene: Research Funding, Speakers Bureau. Sweet:BMS: Honoraria; Astellas: Consultancy; Jazz: Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau; Jazz: Speakers Bureau; Phizer: Consultancy; BMS: Honoraria; Celgene: Honoraria, Speakers Bureau; Astellas: Consultancy; Agios: Consultancy; Phizer: Consultancy; Celgene: Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau; Agios: Consultancy. List:Celgene: Research Funding. Komrokji:Novartis: Honoraria, Speakers Bureau; Celgene: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Novartis: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau. Kuykendall:Janssen: Consultancy; Celgene: Honoraria.

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G-protein-coupled receptors regulate autophagy by ZBTB16-mediated ubiquitination and proteasomal degradation of Atg14L

eLife ◽

10.7554/elife.06734 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 44

Author(s):

Tao Zhang ◽

Kangyun Dong ◽

Wei Liang ◽

Daichao Xu ◽

Hongguang Xia ◽

...

Keyword(s):

Mouse Model ◽

Neurodegenerative Diseases ◽

G Protein ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

G Protein Coupled Receptors ◽

Misfolded Proteins ◽

Ubiquitin Ligase Complex ◽

Wide Range ◽

G Protein Coupled

Autophagy is an important intracellular catabolic mechanism involved in the removal of misfolded proteins. Atg14L, the mammalian ortholog of Atg14 in yeast and a critical regulator of autophagy, mediates the production PtdIns3P to initiate the formation of autophagosomes. However, it is not clear how Atg14L is regulated. In this study, we demonstrate that ubiquitination and degradation of Atg14L is controlled by ZBTB16-Cullin3-Roc1 E3 ubiquitin ligase complex. Furthermore, we show that a wide range of G-protein-coupled receptor (GPCR) ligands and agonists regulate the levels of Atg14L through ZBTB16. In addition, we show that the activation of autophagy by pharmacological inhibition of GPCR reduces the accumulation of misfolded proteins and protects against behavior dysfunction in a mouse model of Huntington's disease. Our study demonstrates a common molecular mechanism by which the activation of GPCRs leads to the suppression of autophagy and a pharmacological strategy to activate autophagy in the CNS for the treatment of neurodegenerative diseases.

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