How do peripheral proteins get to the endoplasmic reticulum surface in yeast?

The Endoplasmic Reticulum (ER) is the entry site into the secretory pathway and is the destination for ~1/3 of yeast proteins that function in the endomembrane system. Mechanisms for targeting transmembrane and luminal proteins to the ER are well-studied. However, it is yet unknown how peripheral proteins reach the ER membrane outer leaflet. We performed a systematic high-content screen in yeast to find factors that affect peripheral ER protein targeting. As our model substrate we used the amphipathic helix-B (HB) from the Brome Mosaic Virus (BMV) protein 1a fused to GFP. HB-GFP has been shown to target specifically to the ER surface. We integrated it into the yeast deletion library to uncover which deletions affect its targeting capacity. We found 137 hits yet none seemed likely to form a new targeting pathway. Rather, our results suggest that membrane lipid composition plays a central role in targeting specificity. We are now verifying these findings on endogenous ER peripheral proteins. By combining molecular dynamics simulations alongside experimental approaches, we aim to unravel the exact membrane characteristics that distinguish the ER from other organelles for binding. Understanding how peripheral ER proteins target correctly is essential for a true comprehension of secretory pathway functions.