Variant-specific interaction of kinectin 1 with the multi-tRNA synthetase complex regulates ER sheet organization

Kinectin 1 (KTN1) is an integral endoplasmic reticulum (ER) sheet protein involved in ER organization and translation elongation. Alternative splicing introduces sequence diversity into the cytosolic region of KTN1; however, the functional significance of these variants has remained unclear. Here, we show that the multi-tRNA synthetase complex (MSC), composed of eight aminoacyl-tRNA synthetases and three nonenzymatic proteins, specifically interacts with KTN1 in a manner dependent on the alternative exon V2 and glutamine-tRNA synthetase (QARS). Using V2 exon-specific knockout cells and KTN1 knockout cells with variant-specific rescue, we found that the V2 exon is required for KTN1-mediated recruitment of the MSC to the ER, where it facilitates the formation of rough ER stacks. These findings define a variant-specific role for KTN1 in anchoring the MSC to the ER and suggest that this interaction underlies a noncanonical function of the MSC in regulating ER sheet organization.