Subproject A06: Specificity of the GET pathway for TA protein insertion in A. thaliana
Principal investigator:
Grefen, Christopher,
Zentrum für Molekularbiologie der Pflanzen (ZMBP)
Abteilung Entwicklungsgenetik
Auf der Morgenstelle 32
72076 Tübingen
Phone: 49 7071 29 73230
E-mail: christopher.grefenspam prevention@zmbp.uni-tuebingen.de
Summary:
Tail-anchored (TA) membrane proteins are involved in a range of important cellular processes such as protein translocation, ubiquitination, energy metabolism, and trafficking to name but a few. They are found in the endomembrane system, as well as in the outer membranes of all subcellular organelles. Specific factors have been suggested to determine the target membrane for any given TA protein and range from size, and lipid composition of the membrane to hydrophobicity, and length of the tail-anchor, and/or charge of its adjacent amino acid sequences, respectively. Also, a discrete pathway was identified – the Guided Entry of TA proteins (GET) pathway – which seems responsible for posttranslational insertion of ER targeted TA proteins. Here, cytosolic chaperones receive the nascent TA protein from the ribosome and shuttle it to ER-localised membrane proteins which facilitate insertion. More recent evidence, however, challenges the view of GET as textbook example for TA protein insertion. For example, get knockout strains in yeast are viable under normal growth conditions, and organ-specific knockout in mice does not cause lethal effects. Our recently published work demonstrates that loss of GET function in Arabidopsis does not globally compromise TA protein insertion, merely leading to root hair growth defects in an otherwise normally developing plant. This is surprising, since SNAREs, one of the largest groups of the TA protein family, facilitate membrane fusion. Dysfunction of this vital step in the secretory pathway should severely affect plant growth and viability.
This project aims to dissect the sequence specificity present in TA proteins that determine GET dependency. We would also like to understand what motifs in the cytosolic chaperone AtGET3a execute TA binding, delivery to downstream receptors or enable alternative functions of the protein. Overexpressing AtGET3a in a receptor knockout (Atget1) leads to severely dwarfed plants with reduced fertility and subcellular AtGET3a aggregates. Hence, the second part of this project is dedicated to decoding the localisation and composition of these aggregates which will help to characterise the specificity of GET proteins in ER membrane insertion and/or a more global involvement of the pathway in proteostasis.