KTN-FLAG-IP for identification of KTN1-variant specific binding proteins

The endoplasmic reticulum (ER) forms sheet or tubule organization, and functions in regulating various cellular processes. Kinectin 1 (KTN1), an integral ER sheet protein involved in ER organization and translation elongation, has alternative exons on the cytoplasmic side, but their roles remain elusive. Here we found that the multi-tRNA synthetase complex (MSC), which is composed of eight aminoacyl-tRNA synthetases (ARSs) and three non-enzymatic proteins, specifically interacts with KTN1 depending on the alternative exon V2 in mouse and human cells. Through a combination of KTN1 knockout and rescue using variant-specific KTN1 expression, we show that the V2 exon is essential for MSC recruitment to the ER, where it contributes to the formation of rough ER stack organization. Our study thus identifies a novel role of KTN1 splicing variant in regulating the ER localization of the MSC, which is likely related to ex-translational activities of MSC to regulate ER sheet organization.