title: Inhibition of N-glycan processing modulates the network of EDEM3 interactors.
abstract: We present here data on EDEM3 network of ER resident interactors and the changes induced upon this network by perturbing the early ER N-glycan processing with mannosidase and glucosidase inhibitors. By coupling immunoprecipitation with mass spectrometry we identified EDEM3 interactors and assigned statistical significance to those most abundant ER-residents that might form functional complexes with EDEM3. We further show that this ER interaction network changes in both content and abundance upon treatment with kifunensine (kif) and N-butyldeoxynojirimycin (NB-DNJ) which suggests that when interfering with the N-glycan processing pathway, the functional complexes involving EDEM3 adapt to maintain the cellular homeostasis. In order to increase the scope of EDEM3 network contenders, the set of MS identified species was further supplemented with putative interactors derived from in silico simulations performed with STRING. Finally, the most interesting candidates to this network were further validated by immunoprecipitation coupled with Western Blotting, which strengthened the confidence in the inferred interactions. The data corroborated herein suggest that besides ER residents, EDEM3 interacts also with proteins involved in the ERAD cargo recognition and targeting to degradation translocation into the cytosol, including UBA1 and UBA2 ubiquitinating enzymes. In addition, the results indicate that this network of EDEM3 interactors is highly sensitive to interfering with early ER N-glycan processing.
authors.affiliation: Departament of Bioinformatics and Structural Biochemistry, Institute of Biochemistry of the Romanian Academy, Splaiul Independenţei, nr. 296, 060031, Bucharest 17, Romania. Electronic address: cristian.butnaru@biochim.ro.
authors.forename: Cristian M
authors.initials: CM
authors.lastname: Butnaru
authors.affiliation: Departament of Molecular Cell Biology, Institute of Biochemistry of the Romanian Academy, Splaiul Independenţei, nr. 296, 060031, Bucharest 17, Romania. Electronic address: mari.chiritoiu@biochim.ro.
authors.forename: Marioara B
authors.initials: MB
authors.lastname: Chiritoiu
authors.affiliation: Departament of Molecular Cell Biology, Institute of Biochemistry of the Romanian Academy, Splaiul Independenţei, nr. 296, 060031, Bucharest 17, Romania. Electronic address: gabriela.chiritoiu@biochim.ro.
authors.forename: Gabriela N
authors.initials: GN
authors.lastname: Chiritoiu
authors.affiliation: Departament of Molecular Cell Biology, Institute of Biochemistry of the Romanian Academy, Splaiul Independenţei, nr. 296, 060031, Bucharest 17, Romania. Electronic address: stefana.petrescu@biochim.ro.
authors.forename: Stefana-Maria
authors.initials: SM
authors.lastname: Petrescu
authors.affiliation: Departament of Bioinformatics and Structural Biochemistry, Institute of Biochemistry of the Romanian Academy, Splaiul Independenţei, nr. 296, 060031, Bucharest 17, Romania. Electronic address: andrei.petrescu@biochim.ro.
authors.forename: Andrei-Jose
authors.initials: AJ
authors.lastname: Petrescu
mesh_terms: D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002460:Cell Line; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D008361:Mannosidases / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D054875:Ubiquitination / Q000502:physiology*; D043323:alpha-Mannosidase
chemical_list: D002135:Calcium-Binding Proteins; D011134:Polysaccharides; D008361:Mannosidases; C573113:EDEM3 protein, human; D043323:alpha-Mannosidase
keywords: EDEM3 interactor network; ERAD pathway; Glycosylation; Mass spectrometry; Ubiquitination