TY - Generic T1 - Identification of two pathways mediating protein targeting from ER to lipid droplets [NOTE: a modified final version was published in Nat Cell Biol and is now available] Y1 - 2021 A1 - Jiunn Song A1 - Arda Mizrak A1 - Chia-Wei Lee A1 - Marcelo Cicconet A1 - Zon Weng Lai A1 - Chieh-Han Lu A1 - Mohr, Stephanie E. A1 - Robert V. Farese, Jr A1 - Tobias C. Walther AB - Pathways localizing proteins to their sites of action within a cell are essential for eukaryotic cell organization and function. Although mechanisms of protein targeting to many organelles have been defined, little is known about how proteins, such as key metabolic enzymes, target from the ER to cellular lipid droplets (LDs). Here, we identify two distinct pathways for ER-to-LD (ERTOLD) protein targeting: early ERTOLD, occurring during LD formation, and late ERTOLD, targeting mature LDs after their formation. By using systematic, unbiased approaches, we identified specific membrane-fusion machinery, including regulators, a tether, and SNARE proteins, that are required for late ERTOLD targeting. Components of this fusion machinery localize to LD-ER interfaces and appear to be organized at ER exit sites (ERES) to generate ER-LD membrane bridges. We also identified multiple cargoes for early and late ERTOLD. Collectively, our data provide a new model for how proteins target LDs from the ER. UR - https://www.biorxiv.org/content/10.1101/2021.09.14.460330v1 ER -