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Protein folding, quality control and degradation in the endoplasmic reticulum

English title Protein folding, quality control and degradation in the endoplasmic reticulum
Applicant Molinari Maurizio
Number 184827
Funding scheme Project funding
Research institution Istituto di ricerca in biomedicina (IRB) Facoltà di scienze biomedice
Institution of higher education Università della Svizzera italiana - USI
Main discipline Cellular Biology, Cytology
Start/End 01.05.2019 - 30.04.2023
Approved amount 833'092.00
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Keywords (5)

Protein misfolding diseases; Proteostasis; Protein folding and quality control; Endoplasmic reticulum (ER); ER-associated degradation

Lay Summary (Italian)

Lead
Il reticolo endoplasmatico è un organello delle cellule nucleate che produce proteine, lipidi e oligosaccaridi. Un suo malfunzionamento è legato all'insorgenza di numerose malattie, molte di queste malattie rare, che derivano dall'incapacità di piegare correttamente le proteine. Queste possono venir distrutte causando malattie da "perdita di funzione" o si possono accumulare causando malattie da "acquisizione di funzione tossica"
Lay summary
Con la nostra ricerca cerchiamo di capire come le cellule del nostro corpo producono le nostre proteine. Studiamo in particolare ciò che avviene nel reticolo endoplasmatico, un organello intracellulare che produce circa il 30% del proteoma Eucariota. Tutte le proteine che devono essere secrete dalle cellule, quelle che devono essere trasportate alla membrana cellulare e nelle membrane e lume degli organelli delle vie secretorie e endocitiche vengono prodotte nel reticolo endoplasmatico. Con l'aiuto di chaperoni molecolari e enzimi, le proteine di nuova sintesi assumono la struttura nativa all'interno del reticolo prima di essere trasportate al sito intra- o extra-cellulare dove eserciteranno le loro funzioni. Il ripiegamento proteico è inefficiente e un sofisticato sistema controlla la qualita dei prodotti genici. Quell che non assumono la struttura corretta vengono distrutti. Nell'impossibilità di farlo, le cellule possono accumularli in aggregati che possono diventare tossici. Sia una degradazione eccessiva, che un accumulo eccessivo di proteine difettose possono essere dannosi alla vita delle nostre cellule e tessuti.
Capire i meccanismi che controllano la qualità dell'espressione genica permetterà di identificare bersagli terapeutici e di intervenire per alleviare i sintomi o curare malattie causate da problemi nella traslazione del materiale genetico in prodotti proteici.
Direct link to Lay Summary Last update: 30.07.2019

Responsible applicant and co-applicants

Employees

Publications

Publication
Quantitative and Time-Resolved Monitoring of Organelle and Protein Delivery to the Lysosome with A Tandem Fluorescent Halo-GFP reporter
Rudinskiy M., Bergmann T.J., Molinari M. (2022), Quantitative and Time-Resolved Monitoring of Organelle and Protein Delivery to the Lysosome with A Tandem Fluorescent Halo-GFP reporter, in Molecular Biology of the Cell.
N‐glycan processing selects ERAD‐resistant misfolded proteins for ER‐to‐lysosome‐associated degradation
Fregno Ilaria, Fasana Elisa, Soldà Tatiana, Galli Carmela, Molinari Maurizio (2021), N‐glycan processing selects ERAD‐resistant misfolded proteins for ER‐to‐lysosome‐associated degradation, in The EMBO Journal, 40(15), e107240.
Cellular Biology of the Endoplasmic Reticulum
Loi Marisa, Marazza Alessandro, Molinari Maurizio (2021), Cellular Biology of the Endoplasmic Reticulum, Springer International Publishing, Cham.
ER-phagy responses in yeast, plants, and mammalian cells and their crosstalk with UPR and ERAD
Molinari Maurizio (2021), ER-phagy responses in yeast, plants, and mammalian cells and their crosstalk with UPR and ERAD, in Developmental Cell, 56(7), 949-966.
Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition) 1
Klionsky Daniel J., Abdel-Aziz Amal Kamal, Abdelfatah Sara, Abdellatif Mahmoud, Abdoli Asghar, Abel Steffen, Abeliovich Hagai, Abildgaard Marie H., Abudu Yakubu Princely, Acevedo-Arozena Abraham, Adamopoulos Iannis E., Adeli Khosrow, Adolph Timon E., Adornetto Annagrazia, Aflaki Elma, Agam Galila, Agarwal Anupam, Aggarwal Bharat B., Agnello Maria, Agostinis Patrizia, Agrewala Javed N., Agrotis Alexander, Aguilar Patricia V., Ahmad S. Tariq, et al. (2021), Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition) 1, in Autophagy, 17(1), 1-382.
Identification of signal peptide features for substrate specificity in human Sec62/Sec63‐dependent ER protein import
Schorr Stefan, Nguyen Duy, Haßdenteufel Sarah, Nagaraj Nagarjuna, Cavalié Adolfo, Greiner Markus, Weissgerber Petra, Loi Marisa, Paton Adrienne W., Paton James C., Molinari Maurizio, Förster Friedrich, Dudek Johanna, Lang Sven, Helms Volkhard, Zimmermann Richard (2020), Identification of signal peptide features for substrate specificity in human Sec62/Sec63‐dependent ER protein import, in The FEBS Journal, 287(21), 4612-4640.
Thioredoxin-Related Transmembrane Proteins: TMX1 and Little Brothers TMX2, TMX3, TMX4 and TMX5
GuerraConcetta, MolinariMaurizio (2020), Thioredoxin-Related Transmembrane Proteins: TMX1 and Little Brothers TMX2, TMX3, TMX4 and TMX5, in Cells, 9(9), 2000.
Deep learning approach for quantification of organelles and misfolded polypeptide delivery within degradative compartments
Morone Diego, Marazza Alessandro, Bergmann Timothy J., Molinari Maurizio (2020), Deep learning approach for quantification of organelles and misfolded polypeptide delivery within degradative compartments, in Molecular Biology of the Cell, 31(14), 1512-1524.
ER-phagy: Eating the Factory
Molinari Maurizio (2020), ER-phagy: Eating the Factory, in Molecular Cell, 78(5), 811-813.
Endoplasmic Reticulum and Lysosomal Quality Control of Four Nonsense Mutants of Iduronate 2-Sulfatase Linked to Hunter's Syndrome
Marazza Alessandro, Galli Carmela, Fasana Elisa, Sgrignani Jacopo, Burda Patricie, Fassi Enrico M.A., Baumgartner Matthias, Cavalli Andrea, Molinari Maurizio (2020), Endoplasmic Reticulum and Lysosomal Quality Control of Four Nonsense Mutants of Iduronate 2-Sulfatase Linked to Hunter's Syndrome, in DNA and Cell Biology, 39(2), 226-234.
Mechanistic insights in recov-ER-phagy: micro-ER-phagy to recover from stress
Loi Marisa, Molinari Maurizio (2020), Mechanistic insights in recov-ER-phagy: micro-ER-phagy to recover from stress, in Autophagy, 16(2), 385-386.
ESCRT-III-driven piecemeal micro-ER-phagy remodels the ER during recovery from ER stress
Loi Marisa, Raimondi Andrea, Morone Diego, Molinari Maurizio (2019), ESCRT-III-driven piecemeal micro-ER-phagy remodels the ER during recovery from ER stress, in Nature Communications, 10(1), 5058-5058.
Proteasomal and lysosomal clearance of faulty secretory proteins: ER-associated degradation (ERAD) and ER-to-lysosome-associated degradation (ERLAD) pathways
Fregno Ilaria, Molinari Maurizio (2019), Proteasomal and lysosomal clearance of faulty secretory proteins: ER-associated degradation (ERAD) and ER-to-lysosome-associated degradation (ERLAD) pathways, in Critical Reviews in Biochemistry and Molecular Biology, 54(2), 153-163.
Schwann cells ER-associated degradation contributes to myelin maintenance in adult nerves and limits demyelination in CMT1B mice
Volpi Vera G., Ferri Cinzia, Fregno Ilaria, Del Carro Ubaldo, Bianchi Francesca, Scapin Cristina, Pettinato Emanuela, Solda Tatiana, Feltri M. Laura, Molinari Maurizio, Wrabetz Lawrence, D’Antonio Maurizio (2019), Schwann cells ER-associated degradation contributes to myelin maintenance in adult nerves and limits demyelination in CMT1B mice, in PLOS Genetics, 15(4), e1008069-e1008069.
A selective ER ‐phagy exerts procollagen quality control via a Calnexin‐ FAM 134B complex
Forrester Alison, De Leonibus Chiara, Grumati Paolo, Fasana Elisa, Piemontese Marilina, Staiano Leopoldo, Fregno Ilaria, Raimondi Andrea, Marazza Alessandro, Bruno Gemma, Iavazzo Maria, Intartaglia Daniela, Seczynska Marta, Anken Eelco, Conte Ivan, De Matteis Maria Antonietta, Dikic Ivan, Molinari Maurizio, Settembre Carmine (2019), A selective ER ‐phagy exerts procollagen quality control via a Calnexin‐ FAM 134B complex, in The EMBO Journal, 38(2), e99847.

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
16th Microbiology and Immunology Introductory Course Individual talk Quantitative steady state and time-resolved analyses of protein and organelle delivery to endolysosomes using Tandem Fluorescent HaloTag-GFP Reporter 19.01.2022 Z Rudinskiy Mikhail;
Simposio Novità nel Campo delle Malattie Metaboliche Talk given at a conference Protein misfolding diseases 09.12.2021 Ascona, Switzerland Molinari Maurizio;
Biotech Biopharm Individual talk Catabolic Control of Endoplasmic Reticulum Homeostasi 06.12.2021 Padua, Italy Fregno Ilaria;
FASEB meeting “The Endoplasmic Reticulum Conference: Structure, Function, and Disease” Talk given at a conference N-glycan processing dictates proteasomal or lysosomal clearance of misfolded proteins from the ER 22.06.2021 E-Meeting, United States of America Fregno Ilaria;
Simposio Inaugurale Centro Malattie Rare Svizzera italiana Talk given at a conference Attualità della Ricerca: Conoscenze sulla deficienza di 1-antitripsina 17.06.2021 Bellinzona, Switzerland Molinari Maurizio;
Virtual live Keystone Symposia eSymposia meeting: “Targeted Protein Degradation: From Small Molecules to Complex Organelles” Talk given at a conference Crosstalk between lysosomal and proteasomal degradation: how N-glycan processing determines the fate of misfolded polypeptides from the ER 07.06.2021 E-meeting, United States of America Fregno Ilaria;
15th Microbiology and Immunology Introductory Course Talk given at a conference Receptor-mediated lysosomal clearance of ER subdomains 11.01.2021 Zurich, Switzerland Kucinska Marika Klaudia;
15th Microbiology and Immunology Introductory Course Individual talk Catabolic Control of Endoplasmic Reticulum Homeostasis 11.01.2021 Zurich, Switzerland Fregno Ilaria;
15th Microbiology and Immunology Introductory Course Poster Tandem Fluorescent HaloTag-GFP Reporter for Quantitative monitoring of protein and organelle delivery to endolysosomes 11.01.2021 Zurich, Switzerland Rudinskiy Mikhail;
GBM/DGZ Fall Conference 2019, Age-Related Human Diseases Special Focus: Autophagy Poster Regulation of the recov-ER-phagy pathway 25.09.2019 Tübingen, Germany Fregno Ilaria;
First LS2 Autophagy Workshop Talk given at a conference ER-phagy, recovER-phagy and ERLAD: the selective turnover of the ER 13.09.2019 Freiburg, Switzerland Molinari Maurizio;
EMBO Workshop Autophagy: From molecular principles to human diseases Poster Mammalian recov-ER-phagy: how does it work? 26.08.2019 Crieff, Great Britain and Northern Ireland Fregno Ilaria;
The 14th ER and Redox Club Meeting Talk given at a conference Eat it right: how recovER-phagy mediates ER clearance 05.08.2019 Herrsching am Ammersee, Germany Molinari Maurizio;
7th Rare Diseases Summer School Poster Endoplasmic reticulum and lysosomal quality control of four nonsense mutants of iduronate 2 sulfatase linked to Hunter’s syndrom 10.07.2019 Kartause Ittingen, Switzerland Marazza Alessandro;
Thirteenth International Calreticulin Workshop Talk given at a conference The role of the Calnexin/Calreticulin cycle in ER-to-lysosome-associated degradation 26.05.2019 Montreal, Canada Molinari Maurizio;
Swiss Proteomic Meeting 2019 – The Annual Meeting of the LS2 Section Proteomics Poster Dissection of cellular responses on perturbation of ER homeostasis using ‘misfoldable’ HaloTag® protein chimeras 23.05.2019 Montreux, Switzerland Guerra Concetta;


Self-organised

Title Date Place
The 15th ER and Redox Club meeting 13.09.2021 Pozzuoli, Italy

Knowledge transfer events

Active participation

Title Type of contribution Date Place Persons involved
Lecture to The Elderly (Uni3, Università terza età) Talk 01.05.2019 Ticino, Switzerland Molinari Maurizio;


Self-organised

Title Date Place
Lectures to High School Students 01.05.2019 High Schools in Ticino, Switzerland

Awards

Title Year
ETH-Zurich Silver Medal for the best PhD 2020

Associated projects

Number Title Start Funding scheme
154421 ER-phagy mechanisms to maintain and restore endoplasmic reticulum homeostasis 01.10.2014 Sinergia

Abstract

Background and rationale-The endoplasmic reticulum (ER) is a biosynthetic organelle of eukaryotic cells. It produces lipids, oligosaccharides and about 40% of the organismal proteome. Protein biogenesis relies on assistance by molecular chaperones, folding enzymes and quality control sensors. These ensure appropriate folding of newly synthesized gene products that are subsequently transported at their intra- or extracellular site of activity. Gene mutations may affect the polypeptide capacity to attain the appropriate structure and function and may enhance the propensity to enter toxic aggregates and polymers. This is linked to an increasing number of so-called protein misfolding diseases, which are characterized by loss-of-function (e.g., on degradation of folding-defective gene products) or gain-of-function phenotypes (e.g., on aggregates or polymers deposition). Folding-defective proteins must be cleared from the ER to maintain the ER capacity to efficiently produce the proteome. Paradoxically, the ER does not contain degradation machineries. Thus, aberrant polypeptides are dislocated across the ER membrane for ER-associated degradation (ERAD) by the ubiquitin proteasome system. Alternatively, misfolded proteins that cannot be dislocated across the ER membrane, or ER subdomains that contain them, are delivered to lysosomal compartments for degradation by ER-to-lysosome-associated degradation (ERLAD) pathways that are poorly understood and whose study is not covered by this SNF allocation. Perturbation of ER homeostasis on unbalanced protein folding, quality control and/or degradation activates unfolded protein responses (UPR) aiming at rescuing or maintaining ER function. On irreversible damages, UPR trigger death programs that eliminate intoxicated cells and can eventually compromise organ homeostasis and organism health. Overall objectives and specific aims-We will characterize the mechanisms that maintain a functional ER by ensuring efficient protein folding and quality control, and dislocation across the ER membrane of misfolded proteins for proteasomal, ER-associated proteins degradation (ERAD). Methods to be used-The pathways regulating protein folding, quality control and degradation will be investigated by following the fate of disease-causing mutant polypeptides, orphan subunits of multimeric proteins, and a series of model polypeptides with specific physico/chemical features generated in our lab to differently challenge the ER (e.g., by inducing or not inducing UPR, by entering or not entering aggregates, by associating or not associating with the membrane). These polypeptides engage various folding, quality control, and ERAD pathways (e.g., by displaying or not displaying oligosaccharides, disulfide bonds, membrane anchors, ...). Expected results and impact-We will characterize the mechanisms that govern protein biogenesis and maintain ER homeostasis. We will study client-specific pathways relying on engagement of dedicated folding, quality control and clearance machineries. Our lab is also interested in elucidating receptor-mediated mechanisms of lysosomal degradation of defective ER material or of defined ER subdomains. The allocation given to our project does not allow to cover the costs of the research on these subjects. Understand protein biogenesis and quality control mechanisms and pathways in molecular details is of crucial importance because their components may be diagnostic and prognostic disease indicators and their modulation is expected to offer therapeutic opportunities in protein misfolding diseases.
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