Investigation on the Role of Sec62 in Membrane Protein Biogenesis

Abstract

Membrane and secretory proteins are synthesized by ribosomes in cytoplasm and targeted to the ER membrane via two pathways

co-translational translocation pathway or post-translational translocation pathway. Membrane proteins are targeted by either the co- or the post-translational translocation pathway and are inserted into the ER membrane by the Sec61 protein conducting channel, which provide a polar passage for polypeptides. During the translocation of polypeptides, Sec61 recognizes a putative transmembrane (TM) segment of about ~20 hydrophobic amino acids and inserts it to the ER membrane through a lateral gate formed by the second and seventh TM segments of the Sec61 channel. Particular for the post-translational translocation pathway is the Sec63/62 protein complex associated with the Sec61 channel. Here, Sec62 acts as a receptor for incoming polypeptides and is found in close proximity to the lateral gate of Sec61. Recently, Reithinger et al (2013) unraveled a novel function of Sec62 in orienting single spanning membrane proteins with a marginally hydrophobic TM segment. However, the function of Sec62 in integration of multi-spanning membrane proteins remained elusive. To investigate the function of Sec62 in integration of multi-spanning membrane proteins, mutations were introduced into several domains of Sec62. When a set of model membrane proteins was expressed in Sec62 mutant strains, the translocation of marginally hydrophobic TM segment of multi spanning membrane proteins into the ER membrane was specifically decreased in Sec62 N-terminal mutant strains. In addition, the interaction between Sec63 and Sec62 was lost in these mutants. Also, reduction of C-terminal translocation of multi spanning membrane proteins was found in Sec62 C-terminal mutant strains as well. Taken together, it is concluded that N-terminus of Sec62 functions by associating Sec62 to the Sec62/63 complex, where Sec62 mediates C-terminal translocation of multi spanning membrane proteins and C-terminus of Sec62 also involves in the process in the ER of Saccharomyces cerevisiae.