Diacidic Motifs in the Carboxyl Terminus Are Required for ER Exit and Translocation to the Plasma Membrane of NKCC2
Abstract
:1. Introduction
2. Results
2.1. Identification of a New Pure ER Exit Signal in NKCC2 COOH-Terminus
2.2. Specificity for the Requirement of 1019DAE1021 Motif in NKCC2 Maturation
2.3. Effect of 1019AAALE1023 and 1019AAALA1023 Mutations on NKCC2 Surface Expression
2.4. Localization of 1019AAALE1023 and 1019AAALA1023 Mutants in the ER
2.5. Mutation of the 1019DAELE1023 Motif Impaired Maturation of the Cotransporter
2.6. Functional Sar1/COPII Machinery Is Required for ER NKCC2 Export
2.7. D1019 and E1021 Residues of the 1019DAE1021 Motif Are Conserved in the COOH-Terminal Tails of All Na-Cl Co-Transporters
3. Discussion
4. Materials and Methods
4.1. Plasmid Constructions and Site Directed Mutagenesis
4.2. Cell Culture
4.3. Protein Preparation and Immunoblotting
4.4. Immunocytochemistry
4.5. Cycloheximide-Chase Assays
4.6. Statistical Analyses
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Bakhos-Douaihy, D.; Seaayfan, E.; Frachon, N.; Demaretz, S.; Kömhoff, M.; Laghmani, K. Diacidic Motifs in the Carboxyl Terminus Are Required for ER Exit and Translocation to the Plasma Membrane of NKCC2. Int. J. Mol. Sci. 2022, 23, 12761. https://doi.org/10.3390/ijms232112761
Bakhos-Douaihy D, Seaayfan E, Frachon N, Demaretz S, Kömhoff M, Laghmani K. Diacidic Motifs in the Carboxyl Terminus Are Required for ER Exit and Translocation to the Plasma Membrane of NKCC2. International Journal of Molecular Sciences. 2022; 23(21):12761. https://doi.org/10.3390/ijms232112761
Chicago/Turabian StyleBakhos-Douaihy, Dalal, Elie Seaayfan, Nadia Frachon, Sylvie Demaretz, Martin Kömhoff, and Kamel Laghmani. 2022. "Diacidic Motifs in the Carboxyl Terminus Are Required for ER Exit and Translocation to the Plasma Membrane of NKCC2" International Journal of Molecular Sciences 23, no. 21: 12761. https://doi.org/10.3390/ijms232112761