Endoplasmic Reticulum (ER)-Targeted, Galectin-Mediated Retrograde Transport by Using a HaloTag Carrier Protein

Abstract

Investigations into metabolic processes within the cell have often relied on genetic methods such as forced expression and knockout or knockdown techniques. An alternative approach would be introducing a molecule into the desired location inside the cell. To translocate compounds from outside cells into the endoplasmic reticulum (ER), we constructed a delivery carrier protein. This comprised N-terminal galectin-1 for cell-surface binding (G1), a protease cleavable sequence (ps), a HaloTag domain for attaching exogenous compounds (Halo), and a C-terminal KDEL sequence for ER retention. Fluorescently labeled G1-ps-Halo-KDEL passed through the Golgi apparatus and reached the ER. By using Man(9)GlcNAc(2)-BODIPY as a cargo compound, the carrier protein was also delivered into the ER with concomitant processing of mannose to Man(5,6), by the ER-resident 1,2-mannosidase. G1-ps-Halo-KDEL might serve as a new type of delivery carrier protein to direct compounds into the ER.