Abstract N-terminal signal peptides (SPs) are traditionally considered as drivers of co-translational translocation of newly synthesised proteins into the endoplasmic reticulum (ER). However, growing evidences suggest that proteins with SPs can also undergo post-translational insertion into the ER membrane after synthesis is complete. Recently, an intermediate third mechanism has been uncovered where proteins with marginally hydrophobic or suboptimal SPs are translocated following an initial delay after translation initiation. Here, we show that this “delayed translocation” allows a temporary exposure of the nascent chain to the cytosolic environment, enabling exoplasmic domain modifications by cytosolic enzymes. We report that programmed death ligand-1 (PD-L1) follows this pathway, featuring a suboptimal SP that exposes its extracellular domain to the cytosol, enabling AMPK-dependent regulation of PD-L1 function. Importantly, optimising the SP of PD-L1 eliminates the cytosolic exposure, disrupting PD-L1’s trafficking and maturation, highlighting the physiological importance of the delayed translocation mechanism.
Serio et al. (Sat,) studied this question.