{Reference Type}: Journal Article
{Title}: A proximity labeling-based orthogonal trap strategy identifies HDAC8 promotes cell motility by modulating cortactin acetylation.
{Author}: Huang Y;Zhai G;Fu Y;Li Y;Zang Y;Lin Y;Zhang K;
{Journal}: Cell Chem Biol
{Volume}: 31
{Issue}: 3
{Year}: 2024 Mar 21
{Factor}: 9.039
{DOI}: 10.1016/j.chembiol.2024.02.003
{Abstract}: It is a challenge for the traditional affinity methods to capture transient interactions of enzyme-post-translational modification (PTM) substrates in vivo. Herein we presented a strategy termed proximity labeling-based orthogonal trap approach (ProLORT), relying upon APEX2-catalysed proximity labeling and an orthogonal trap pipeline as well as quantitative proteomics to directly investigate the transient interactome of enzyme-PTM substrates in living cells. As a proof of concept, ProLORT allows for robust evaluation of a known HDAC8 substrate, histone H3K9ac. By leveraging this approach, we identified numerous of putative acetylated proteins targeted by HDAC8, and further confirmed CTTN as a bona fide substrate in vivo. Next, we demonstrated that HDAC8 facilitates cell motility via deacetylation of CTTN at lysine 144 that attenuates its interaction with F-actin, expanding the underlying regulatory mechanisms of HDAC8. We developed a general strategy to profile the transient enzyme-substrate interactions mediated by PTMs, providing a powerful tool for identifying the spatiotemporal PTM-network regulated by enzymes in living cells.