Dataset Information

Pevonedistat inhibits aberrant protein NEDDylation in Polycystic Liver Diseases.

ABSTRACT: Background: Polycystic liver diseases (PLD) are genetic disorders characterized by progressive growth of numerous liver cysts, causing significant morbidity. Previous studies revealed genes affecting protein biogenesis and more recently, protein SUMOylation, a posttranslational modification (PTM), has been implicated in PLD pathobiology. On the other hand, protein NEDDylation is a newly-characterized PTM, modulating a plethora of biological processes and its dysregulation is associated with development and progression of several human diseases. In this regard, the role of NEDDylation in PLD remains elusive and unraveling its role in the pathogenesis of these genetic disorders could open new avenues for the development of novel treatments in the future. Objective: To explore the role of protein NEDDylation in PLD and its potential therapeutic regulatory value. Methods: Expression and function of NEDDylation, including response to Pevonedistat (first-in-class selective inhibitor of the NEDDylation E1 enzyme NAE1), were assessed in vitro. Proteomic analyses of immunoprecipitated NEDDylated proteins were performed by mass spectrometry.

INSTRUMENT(S): timsTOF Pro

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cholangiocyte, Liver

DISEASE(S): Polycystic Liver Disease

SUBMITTER: Mikel Azkargorta

LAB HEAD: Felix Elortza

PROVIDER: PXD022571 | Pride | 2022-02-16

REPOSITORIES: Pride

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Publications

Inhibition of NAE-dependent protein hyper-NEDDylation in cystic cholangiocytes halts cystogenesis in experimental models of polycystic liver disease.

Lee-Law Pui Y PY Olaizola Paula P Caballero-Camino Francisco J FJ Izquierdo-Sanchez Laura L Rodrigues Pedro M PM Perugorria Maria J MJ Azkargorta Mikel M Elortza Felix F Martinez-Chantar Maria L ML Aspichueta Patricia P Marzioni Marco M Bujanda Luis L Drenth Joost P H JPH Banales Jesus M JM

United European gastroenterology journal 20210726 7

<h4>Background</h4>Polycystic liver diseases (PLDs) are genetic inherited disorders characterized by the progressive growth of numerous intrahepatic biliary cysts, which are the main cause of morbidity. Previous studies revealed that cystic cholangiocytes are characterized by endoplasmic reticulum stress and aberrant posttranslational modification (PTM) of proteins, in particular hyper-SUMOylation, that promote PLD pathobiology. Protein NEDDylation is a newly characterized PTM that modulates a p ...[more]

PMID: 34310849

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Project description:Polycystic liver disease (PLD) is a group of genetic disorders characterized by bile duct dilatation and/or progressive development of multiple intrahepatic fluid-filled biliary cysts (>10), which are the main cause of morbidity. Current surgical and/or pharmacological therapies exert short-term and modest benefits, and thus liver transplantation represents the only curative option. Recently, novel PLD-causative genes, encoding for endoplasmic reticulum (ER)-resident proteins involved in protein biogenesis and transport, were identified. Here, we hypothesized that aberrant proteostasis contributes to PLD pathogenesis, representing a potential therapeutic target. In this regard, ER stress was assessed at transcriptional (qPCR), proteomic (mass spectrometry), morphological (transmission electron microscopy; TEM) and functional (20S proteasome activity) levels in different models of PLD (i.e., human and rat). Moreover, PCK rat (animal model of PLD) was employed to determine the effects of ER stress inhibitors [4-phenylbutyric acid (4-PBA)] and/or inducers [tunicamycin (TM)] in hepatic cytogenesis and, consequently, in the progression of the disease. Likewise, the impact of the aforementioned ER stress modulators was also analyzed on the expression of unfolded protein response (UPR) factors, the 20S proteasome activity, the mechanisms involved in the maintenance of ER proteostasis and the cell fate decisions (i.e., survival/apoptosis). Interestingly, the expression levels of the UPR factors were markedly upregulated in liver tissue from PLD patients and PCK rats, as well as in primary cultures of human and rat cystic cholangiocytes, compared to normal controls. Additionally, cystic cholangiocytes showed altered proteomic profiles, mainly related to proteostasis (i.e., synthesis, folding, trafficking and degradation of nascent proteins), marked enlargement of the ER lumen (by TEM), and hyperactivation of the 20S proteasome. Notably, chronic treatment of PCK rats with 4-PBA decreased liver weight, as well as both liver and cystic volumes, of animals under baseline or TM administration compared to control groups. In vitro, 4-PBA downregulated the expression (mRNA) of UPR effectors, normalized, at least in part, proteomic profiles related to protein synthesis, folding, trafficking and degradation, and reduced the proteasome hyperactivity in cystic cholangiocytes, reducing their hyperproliferation and apoptosis. Conclusion: Restoration of proteostasis in cystic cholangiocytes with 4-PBA halts hepatic cystogenesis, emerging as a novel and promising therapeutic strategy.

Project description:A time course of the effect of overexpressing GPI-PLD on global gene expression in HepG2 cells was compared to control virus. Experiment Overall Design: This experiment compares the global gene expression in HepG2 cells that were either transduced with an adenovirus expressing GPI-phospholipase D or an adenovirus expressing beta galactosidase as a control. Treatment was for 6 or 12 hrs. Four replicates for each condition were used. 10 micrograms of total RNA was assayed per genechip using standard Affymetrix protocols. CONTEXT: Recent studies demonstrated that de novo lipogenesis is increased in patients with nonalcoholic fatty liver disease (NAFLD). Patients with NAFLD also have plasma lipid abnormalities. These lipid abnormalities may in part be related to insulin resistance, which is common in patients with NAFLD. Insulin resistance is associated with alterations in proteins involved in lipid metabolism including glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD), which is involved in triglyceride metabolism. OBJECTIVE: The objective of the study was to determine whether alterations in serum and hepatic levels of GPI-PLD occur in patients with NAFLD. DESIGN AND PATIENTS: We examined the following: 1) levels of serum GPI-PLD in nondiabetics with nonalcoholic steatohepatitis, compared with matched controls; 2) hepatic expression of GPI-PLD mRNA in patients with normal liver or NAFLD; and 3) effect of overexpressing GPI-PLD vs. beta-galactosidase (control) on global gene expression in a human hepatoma cell line. RESULTS: The serum levels of GPI-PLD were significantly higher in patients with nonalcoholic steatohepatitis than in matched controls (119 +/- 24 vs.105 +/- 15 microg/ml, P = 0.047). The hepatic expression of GPI-PLD mRNA was increased nearly 3-fold in NAFLD patients, compared with patients with normal liver (3.1 +/- 2.6 vs. 1.1 +/- 1.0 arbitrary units per microgram total RNA, P = 0.026). Finally, overexpressing GPI-PLD was associated with an increase in de novo lipogenesis genes. CONCLUSIONS: Patients with NAFLD have elevated serum levels and hepatic expression of GPI-PLD, and its overexpression in vitro is associated with increased expression of de novo lipogenesis genes. These results suggest that GPI-PLD may play a role in the pathogenesis of NAFLD and/or its metabolic features and warrants further investigation.

Dataset Information

Pevonedistat inhibits aberrant protein NEDDylation in Polycystic Liver Diseases.

Publications

Inhibition of NAE-dependent protein hyper-NEDDylation in cystic cholangiocytes halts cystogenesis in experimental models of polycystic liver disease.

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