Genetically encoded fluorescent tools: Shining a little light on ER-to-Golgi transport

Abstract

Since the first fluorescent proteins (FPs) were identified and isolated over fifty years ago, FPs have become commonplace yet indispensable tools for studying the constitutive secretory pathway in live cells. At the same time, genetically encoded chemical tags have provided a new use for much older fluorescent dyes. Innovation has also produced several specialized methods to allow synchronous release of cargo proteins from the endoplasmic reticulum (ER), enabling precise characterization of sequential trafficking steps in the secretory pathway. Without the constant innovation of the researchers who design these tools to control, image, and quantitate protein secretion, major discoveries about ER-to-Golgi transport and later stages of the constitutive secretory pathway would not have been possible. We review many of the tools and tricks, some 25 years old and others brand new, that have been successfully implemented to study ER-to-Golgi transport in intact and living cells.

Keywords: ER to Golgi transport; Endoplasmic reticulum; Fluorescent imaging; Fluorescent proteins; Golgi apparatus; Secretory pathway; Transport assay; Transport vesicle; Vesicle trafficking; Vesicle transport.

Figure 1.. Methods for synchronous release of cargo proteins from the ER to study ER-to-Golgi and subsequent trafficking steps.

(A) Temperature-shift methods established for collagen and VSV-G_tsO45 transport. In addition, common temperature blocks for synchronizing any cargoes at the ERGIC or TGN are described in the text. (B) Methods based upon the ARIAD/iDimerize system for controlled aggregation of cargoes in the ER. (C) The RUSH system and several of the readily available cargoes for ER to plasma membrane trafficking. (D) Light-activated trafficking methods established for wt VSV-G and transferrin receptor. See text for details and references.