Endoplasmic reticulum membrane receptors of the GET pathway are conserved throughout eukaryotes

• Published in: Proceedings of the National Academy of Sciences - PNAS 2021-01, Vol.118 (1), p.1-9
• Main Authors: Asseck, Lisa Yasmin, Mehlhorn, Dietmar Gerald, Monroy, Jhon Rivera, Ricardi, Martiniano Maria, Breuninger, Holger, Wallmeroth, Niklas, Berendzen, Kenneth Wayne, Nowrousian, Minou, Xing, Shuping, Schwappach, Blanche, Bayer, Martin, Grefen, Christopher
• Format: Article
• Language: English
• Subjects: Adaptor Proteins, Vesicular Transport - genetics; Adaptor Proteins, Vesicular Transport - metabolism; Adenosine triphosphatase; Apoptosis; Arabidopsis; Arabidopsis - genetics; Arabidopsis - metabolism; Arabidopsis Proteins - genetics; Arabidopsis Proteins - metabolism; Biological Sciences; Cell Membrane - metabolism; Conserved sequence; Cytosol; Cytosol - metabolism; Ectopic expression; Endoplasmic reticulum; Endoplasmic Reticulum - metabolism; Eukaryotes; Homology; Immunoprecipitation; Insertion; Intracellular Membranes - metabolism; Mammals; Mass spectrometry; Mass spectroscopy; Membrane proteins; Membrane Proteins - metabolism; Membranes; Phenotype; Phenotypes; Protein Transport; Proteins; Receptors; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Translocation; Yeast; Yeasts
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.2017636118

Type II tail-anchored (TA) membrane proteins are involved in diverse cellular processes, including protein translocation, vesicle trafficking, and apoptosis. They are characterized by a single C-terminal transmembrane domain that mediates posttranslational targeting and insertion into the endoplasmic reticulum (ER) via the Guided-Entry of TA proteins (GET) pathway. The GET system was originally described in mammals and yeast but was recently shown to be partially conserved in other eukaryotes, such as higher plants. A newly synthesized TA protein is shielded from the cytosol by a pretargeting complex and an ATPase that delivers the protein to the ER, where membrane receptors (Get1/WRB and Get2/CAML) facilitate insertion. In the model plant Arabidopsis thaliana, most components of the pathway were identified through in silico sequence comparison, however, a functional homolog of the coreceptor Get2/CAML remained elusive. We performed immunoprecipitation- mass spectrometry analysis to detect in vivo interactors of AtGET1 and identified a membrane protein of unknown function with low sequence homology but high structural homology to both yeast Get2 and mammalian CAML. The protein localizes to the ER membrane, coexpresses with AtGET1, and binds to Arabidopsis GET pathway components. While loss-of-function lines phenocopy the stunted root hair phenotype of other Atget lines, its heterologous expression together with the coreceptor AtGET1 rescues growth defects of Δget1get2 yeast. Ectopic expression of the cytosolic, positively charged N terminus is sufficient to block TA protein insertion in vitro. Our results collectively confirm that we have identified a plant-specific GET2 in Arabidopsis, and its sequence allows the analysis of cross-kingdom pathway conservation.