EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning
EDEM3 recognizes and directs misfolded proteins to the ER-associated protein degradation (ERAD) process. EDEM3 was predicted to act as lectin or as a mannosidase because of its homology with the GH47 catalytic domain of the Man1B1, but the contribution of the other regions remained unresolved. Here,...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-02-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/22/4/2172 |
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| author | Georgiana Manica Simona Ghenea Cristian V. A. Munteanu Eliza C. Martin Cristian Butnaru Marius Surleac Gabriela N. Chiritoiu Petruta R. Alexandru Andrei-Jose Petrescu Stefana M. Petrescu |
| author_facet | Georgiana Manica Simona Ghenea Cristian V. A. Munteanu Eliza C. Martin Cristian Butnaru Marius Surleac Gabriela N. Chiritoiu Petruta R. Alexandru Andrei-Jose Petrescu Stefana M. Petrescu |
| author_sort | Georgiana Manica |
| collection | DOAJ |
| description | EDEM3 recognizes and directs misfolded proteins to the ER-associated protein degradation (ERAD) process. EDEM3 was predicted to act as lectin or as a mannosidase because of its homology with the GH47 catalytic domain of the Man1B1, but the contribution of the other regions remained unresolved. Here, we dissect the molecular determinants governing EDEM3 function and its cellular interactions. LC/MS analysis indicates very few stable ER interactors, suggesting EDEM3 availability for transient substrate interactions. Sequence analysis reveals that EDEM3 consists of four consecutive modules defined as GH47, intermediate (IMD), protease-associated (PA), and intrinsically disordered (IDD) domain. Using an EDEM3 knock-out cell line, we expressed EDEM3 and domain deletion mutants to address EDEM3 function. We find that the mannosidase domain provides substrate binding even in the absence of mannose trimming and requires the IMD domain for folding. The PA and IDD domains deletions do not impair the trimming, but specifically modulate the turnover of two misfolded proteins, NHK and the soluble tyrosinase mutant. Hence, we demonstrate that EDEM3 provides a unique ERAD timing to misfolded glycoproteins, not only by its mannose trimming activity, but also by the positive and negative feedback modulated by the protease-associated and intrinsically disordered domain, respectively. |
| first_indexed | 2024-03-09T00:38:34Z |
| format | Article |
| id | doaj.art-813729637e174c028af2ec0c5de414f7 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-09T00:38:34Z |
| publishDate | 2021-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-813729637e174c028af2ec0c5de414f72023-12-11T17:58:12ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-02-01224217210.3390/ijms22042172EDEM3 Domains Cooperate to Perform Its Overall Cell FunctioningGeorgiana Manica0Simona Ghenea1Cristian V. A. Munteanu2Eliza C. Martin3Cristian Butnaru4Marius Surleac5Gabriela N. Chiritoiu6Petruta R. Alexandru7Andrei-Jose Petrescu8Stefana M. Petrescu9Department of Molecular Cell Biology, Institute of Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Molecular Cell Biology, Institute of Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Bioinformatics and Structural Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Bioinformatics and Structural Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Bioinformatics and Structural Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Bioinformatics and Structural Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Molecular Cell Biology, Institute of Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Molecular Cell Biology, Institute of Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Bioinformatics and Structural Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaDepartment of Molecular Cell Biology, Institute of Biochemistry, Splaiul Independentei 296, 060031 Bucharest 17, RomaniaEDEM3 recognizes and directs misfolded proteins to the ER-associated protein degradation (ERAD) process. EDEM3 was predicted to act as lectin or as a mannosidase because of its homology with the GH47 catalytic domain of the Man1B1, but the contribution of the other regions remained unresolved. Here, we dissect the molecular determinants governing EDEM3 function and its cellular interactions. LC/MS analysis indicates very few stable ER interactors, suggesting EDEM3 availability for transient substrate interactions. Sequence analysis reveals that EDEM3 consists of four consecutive modules defined as GH47, intermediate (IMD), protease-associated (PA), and intrinsically disordered (IDD) domain. Using an EDEM3 knock-out cell line, we expressed EDEM3 and domain deletion mutants to address EDEM3 function. We find that the mannosidase domain provides substrate binding even in the absence of mannose trimming and requires the IMD domain for folding. The PA and IDD domains deletions do not impair the trimming, but specifically modulate the turnover of two misfolded proteins, NHK and the soluble tyrosinase mutant. Hence, we demonstrate that EDEM3 provides a unique ERAD timing to misfolded glycoproteins, not only by its mannose trimming activity, but also by the positive and negative feedback modulated by the protease-associated and intrinsically disordered domain, respectively.https://www.mdpi.com/1422-0067/22/4/2172EDEM3ERADNHKtyrosinasemass spectrometryER mannosidases |
| spellingShingle | Georgiana Manica Simona Ghenea Cristian V. A. Munteanu Eliza C. Martin Cristian Butnaru Marius Surleac Gabriela N. Chiritoiu Petruta R. Alexandru Andrei-Jose Petrescu Stefana M. Petrescu EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning International Journal of Molecular Sciences EDEM3 ERAD NHK tyrosinase mass spectrometry ER mannosidases |
| title | EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning |
| title_full | EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning |
| title_fullStr | EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning |
| title_full_unstemmed | EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning |
| title_short | EDEM3 Domains Cooperate to Perform Its Overall Cell Functioning |
| title_sort | edem3 domains cooperate to perform its overall cell functioning |
| topic | EDEM3 ERAD NHK tyrosinase mass spectrometry ER mannosidases |
| url | https://www.mdpi.com/1422-0067/22/4/2172 |
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