Membrane lipids couple synaptotagmin to SNARE-mediated granule fusion in insulin-secreting cells.

Insulin secretion depends on the Ca2+-regulated fusion of granules with the plasma membrane. A recent model of Ca2+-triggered exocytosis in secretory cells proposes that lipids in the plasma membrane couple the calcium sensor Syt1 to the membrane fusion machinery (Kiessling et al., 2018). Specifically, Ca2+-mediated binding of Syt1's C2 domains to the cell membrane shifts the membrane-anchored SNARE syntaxin-1a to a more fusogenic conformation, straightening its juxtamembrane linker. To test this model in live cells and extend it to insulin secretion, we enriched INS1 cells with a panel of lipids with different acyl chain compositions. Fluorescence lifetime measurements demonstrate that cells with more disordered membranes show an increase in fusion efficiency, and vice versa. Experiments with granules purified from INS1 cells and recombinant SNARE proteins reconstituted in supported membranes confirmed that lipid acyl chain composition determines SNARE conformation and that lipid disordering correlates with increased fusion. Addition of Syt1's C2AB domains significantly decreased lipid order in target membranes and increased SNARE-mediated fusion probability. Strikingly, Syt's action on both fusion and lipid order could be partially bypassed by artificially increasing unsaturated phosphatidylserines in the target membrane. Thus, plasma membrane lipids actively participate in coupling Ca2+/synaptotagmin-sensing to the SNARE fusion machinery in cells.

Erythroid Differentiation Dependent Interaction of VPS13A with XK at the Plasma Membrane of K562 Cells.

Mutations of the bridge-like lipid transport protein VPS13A and the lipid scramblase XK result in Chorea Acanthocytosis (ChAc) and McLeod syndrome (MLS), respectively, two similar conditions involving neurodegeneration and deformed erythrocytes (acanthocytes). VPS13A binds XK, suggesting a model in which VPS13A forms a lipid transport bridge between the endoplasmic reticulum (ER) and the plasma membrane (PM), where XK resides. However, studies of VPS13A in HeLa and COS7 cells showed that this protein localizes primarily at contacts of the ER with mitochondria. Overexpression of XK in these cells redistributed VPS13A to the biosynthetic XK pool in the ER but not to PM-localized XK. Colocalization of VPS13A with XK at the PM was only observed if overexpressed XK harbored mutations that disengaged its VPS13A-binding site from an intramolecular interaction. As the acanthocytosis phenotype of ChAc and MLS suggests a role of the two proteins in cells of the erythroid lineage, we explored their localization in K562 cells, which differentiate into erythroblasts upon hemin addition. When tagged VPS13A was overexpressed in hemin-treated K562 cells, robust formation of ER-PM contacts positive for VPS13A was observed and their formation was abolished in XK KO cells. ER-PM contacts positive for VPS13A were seldom observed in undifferentiated K562 cells, despite the presence of XK in these cells at concentrations similar to those observed after differentiation. These findings reveal that the interaction of VPS13A with XK at ER-PM contacts requires a permissive state which depends upon cell type and/or functional state of the cell.

Collagen XIX is required for pheromone recognition and glutamatergic synapse formation in mouse accessory olfactory bulb.

In mammals, the accessory olfactory bulb (AOB) receives input from vomeronasal sensory neurons (VSN) which detect pheromones, chemical cues released by animals to regulate the physiology or behaviors of other animals of the same species. Cytoarchitecturally, cells within the AOB are segregated into a glomerular layer (GL), mitral cell layer (MCL), and granule cell layer (GCL). While the cells and circuitry of these layers has been well studied, the molecular mechanism underlying the assembly of such circuitry in the mouse AOB remains unclear. With the goal of identifying synaptogenic mechanisms in AOB, our attention was drawn to Collagen XIX, a non-fibrillar collagen generated by neurons in the mammalian telencephalon that has previously been shown to regulate the assembly of synapses. Here, we used both a targeted mouse mutant that lacks Collagen XIX globally and a conditional allele allowing for cell-specific deletion of this collagen to test if the loss of Collagen XIX causes impaired synaptogenesis in the mouse AOB. These analyses not only revealed defects in excitatory synapse distribution in these Collagen XIX-deficient mutants, but also showed that these mutant mice exhibit altered behavioral responses to pheromones. Although this collagen has been demonstrated to play synaptogenic roles in the telencephalon, those roles are at perisomatic inhibitory synapses, results here are the first to demonstrate the function of this unconventional collagen in glutamatergic synapse formation.