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Abstract

During membrane protein biogenesis, cells need to detect and degrade faulty proteins. Despite a key role in cellular homeostasis and human diseases, little is known about the underlying mechanisms. In recent years, few endoplasmic reticulum (ER)-resident proteases have been linked to quality control by cleaving their clients and thereby facilitating membrane extraction and degradation via the ER-associated degradation (ERAD) pathway. The major ER-resident protease in mammalian cells is the signal peptidase complex (SPC), a tetra subunit complex discovered in the 1970s to be responsible for the removal of signal sequences from ER-targeted and secretory proteins. Until now, this was thought to be the only function of the SPC besides a few studies reporting a role in the maturation of viral polyproteins. In this work, I show that the SPC also acts as a membrane protein quality control factor. First, through proteome-wide computational analyses, I identified approximately 1500 membrane proteins containing SPC cryptic cleavage sites after N-terminal and internal type-II oriented transmembrane domains (TMDs). I then validated SPC cleavage for several candidate substrates (Cx32, Cx26, Cx30.3, PMP22, iRhom2 and Hrd1) and revealed that SPC cleavage relies on the accessory subunit SPCS1 as recognition factor to discern between signal sequences and TMDs. Moreover, I show that the SPC cleaves membrane proteins when they fail to fold properly or assemble correctly into their native complexes, thus exposing cryptic cleavage sites. I also show that this SPC cleavage mechanism cooperates with the ERAD pathway to help maintain a functional membrane proteome and confers a fitness advantage to cells exposed to ER stress. Finally, I report first data on the possible role of the SPC in controlling protein abundance beyond its quality control function. Overall, this thesis characterises a novel function of the SPC, expanding its substrate spectrum, extending the knowledge on the essential cellular functions performed by this protease and laying the foundations for future work at the organismal level in quality control-related diseases and beyond.