Journal: The EMBO Journal

Article Title: Mutations in the kinesin KIF12 promote MASH in humans and mice by disrupting lipogenic enzyme turnover

doi: 10.1038/s44318-025-00366-8

Figure Lengend Snippet: ( A ) Immunoblotting of BioID assay precipitates in HepG2 cells stably transduced with KIF12-PRD-BioID2 (PRD) and BioID2 alone (NC) constructs. ( B ) GST pulldown assay of mouse liver lysates (Input) against KIF12-PRD-GST (PRD) and GST-alone (NC), shown by ACC1 immunoblotting (upper) and CBB staining (lower). Note that GST-PRD specifically pulled down ACC1. ( C – E ) Fluorescence images of a tag-proximity ligation assay in HepG2 cells being transduced with mCit-KIF12-FL, mCit-KIF12-PRD, and mCit alone (NC), respectively, which were proximity-labeled by an anti-GFP antibody and anti-ACC1 or anti-PC antibodies ( C ), with statistics ( D and E ). Scale bars, 10 μm. Error bars, mean ± SEM. Welch’s t test. Biological replicates, individual cells. Arrows, protein interaction signals.

Article Snippet: For generating the pPRD-BioID2 expression vector, cDNA encoding the KIF12-PRD was amplified with the N-BAM-ATG and C-BAM-NoStop primers, and ligated with the MCS-BioID2-HA vector (kindly provided by Dr. Kyle Roux through Addgene, #74224) (Kim et al, ) at the BamHI restriction site, and the directions of the inserts were verified by Sanger sequencing.

Techniques: Western Blot, Stable Transfection, Transduction, Construct, GST Pulldown Assay, Staining, Fluorescence, Proximity Ligation Assay, Labeling