Resistance to Inhibitors of Cholinesterase (Ric)-8A and Gα(i) Contribute to Cytokinesis Abscission by Controlling Vacuolar Protein-Sorting (Vps)34 Activity

Resistance to inhibitors of cholinesterase (Ric)-8A is a guanine nucleotide exchange factor for Gα(i), Gα(q), and Gα(12/13), which is implicated in cell signaling and as a molecular chaperone required for the initial association of nascent Gα subunits with cellular membranes. Ric-8A, Gα(i) subunits, and their regulators are localized at the midbody prior to abscission and linked to the final stages of cell division. Here, we identify a molecular mechanism by which Ric-8A affects cytokinesis and abscission by controlling Vps34 activity. We showed that Ric-8A protein expression is post-transcriptionally controlled during the cell cycle reaching its maximum levels at mitosis. A FRET biosensor created to measure conformational changes in Ric-8A by FLIM (Fluorescence Lifetime Imaging Microscopy) revealed that Ric-8A was in a close-state during mitosis and particularly so at cytokinesis. Lowering Ric-8A expression delayed the abscission time of dividing cells, which correlated with increased intercellular bridge length and multinucleation. During cytokinesis, Ric-8A co-localized with Vps34 at the midbody along with Gα(i) and LGN, where these proteins functioned to regulate Vps34 phosphatidylinositol 3-kinase activity.