Acyl-CoA-binding protein, Acb1p, is required for normal vacuole function and ceramide synthesis in Saccharomyces cerevisiae.

In the present study, we show that depletion of acyl-CoA-binding protein, Acb1p, in yeast affects ceramide levels, protein trafficking, vacuole fusion and structure. Vacuoles in Acb1p-depleted cells are multi-lobed, contain significantly less of the SNAREs (soluble N -ethylmaleimide-sensitive fusion protein attachment protein receptors) Nyv1p, Vam3p and Vti1p, and are unable to fuse in vitro. Mass spectrometric analysis revealed a dramatic reduction in the content of ceramides in whole-cell lipids and in vacuoles isolated from Acb1p-depleted cells. Maturation of yeast aminopeptidase I and carboxypeptidase Y is slightly delayed in Acb1p-depleted cells, whereas the maturation of alkaline phosphatase and Gas1p is unaffected. The fact that Gas1p maturation is unaffected by Acb1p depletion, despite the lowered ceramide content in these cells, indicates that ceramide synthesis in yeast could be compartmentalized. We suggest that the reduced ceramide synthesis in Acb1p-depleted cells leads to severely altered vacuole morphology, perturbed vacuole assembly and strong inhibition of homotypic vacuole fusion

Antibody-mediated targeting of the Orai1 calcium channel inhibits T cell function.

Despite the attractiveness of ion channels as therapeutic targets, there are no examples of monoclonal antibodies directed against ion channels in clinical development. Antibody-mediated inhibition of ion channels could offer a directed, specific therapeutic approach. To investigate the potential of inhibiting ion channel function with an antibody, we focused on Orai1, the pore subunit of the calcium channel responsible for store-operated calcium entry (SOCE) in T cells. Effector T cells are key drivers of autoimmune disease pathogenesis and calcium signaling is essential for T cell activation, proliferation, and cytokine production. We show here the generation of a specific anti-human Orai1 monoclonal antibody (mAb) against an extracellular loop of the plasma membrane-spanning protein. The anti-Orai1 mAb binds native Orai1 on lymphocytes and leads to cellular internalization of the channel. As a result, T cell proliferation, and cytokine production is inhibited in vitro. In vivo, anti-Orai1 mAb is efficacious in a human T cell-mediated graft-versus host disease (GvHD) mouse model. This study demonstrates the feasibility of antibody-mediated inhibition of Orai1 function and, more broadly, reveals the possibility of targeting ion channels with biologics for the treatment of autoimmunity and other diseases