title: Targeting and membrane insertion into the endoplasmic reticulum membrane of Saccharomyces cerevisiae essential protein Rot1.
abstract: Rot1 is an essential yeast protein that has been related to cell wall biosynthesis, actin cytoskeleton dynamics and protein folding. Rot1 is an N-glycosylated protein anchored to the nuclear envelope-endoplasmic reticulum (ER) membrane by a transmembrane domain at its C-terminal end. Rot1 is translocated to the ER by a post-translational mechanism. Here, we investigate the protein domain required to target and translocate Rot1 to the ER membrane. We found that several deletions of the N-terminal region of Rot1 prevented neither membrane targeting nor the insertion of this protein. Interestingly, we obtained the same results when different truncated forms in the C-terminal transmembrane domain were analyzed, suggesting the presence of an internal topogenic element that is capable of translocating Rot1 to the ER. To identify this sequence, we generated a combination of N- and C-terminal deletion mutants of Rot1 and we investigated their insertion into the membrane. The results show that two regions, amino acids 26-60 and 200-228, are involved in the post-translational translocation of Rot1 across the ER membrane.
authors.affiliation: Departament de Bioquímica i Biologia Molecular, Universitat de València, València, Spain.
authors.forename: María Angeles
authors.initials: MA
authors.lastname: Juanes
authors.forename: Carlos Andrés
authors.initials: CA
authors.lastname: Martínez-Garay
authors.forename: Juan Carlos
authors.initials: JC
authors.lastname: Igual
authors.forename: María Carmen
authors.initials: MC
authors.lastname: Bañó
mesh_terms: D004721:Endoplasmic Reticulum / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D017384:Sequence Deletion
chemical_list: D008565:Membrane Proteins; D018832:Molecular Chaperones; C523667:ROT1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins