The lipid distribution of plasma membranes of eukaryotic cells is asymmetric and phospholipid scramblases disrupt this asymmetry by mediating the rapid, nonselective transport of lipids down their concentration gradients. As a result, phosphatidylserine is exposed to the outer leaflet of membrane, an important step in extracellular signaling networks controlling processes such as apoptosis, blood coagulation, membrane fusion and repair. Several TMEM16 family members have been identified as Ca2+-activated scramblases, but the mechanisms underlying their Ca2+-dependent gating and their effects on the surrounding lipid bilayer remain poorly understood. Here, we describe three high-resolution cryo-electron microscopy structures of a fungal scramblase from Aspergillus fumigatus, afTMEM16, reconstituted in lipid nanodiscs. These structures reveal that Ca2+-dependent activation of the scramblase entails global rearrangement of the transmembrane and cytosolic domains. These structures, together with functional experiments, suggest that activation of the protein thins the membrane near the transport pathway to facilitate rapid transbilayer lipid movement.

Ceramides are sphingolipids that modulate a variety of cellular processes via 2 major mechanisms: functioning as second messengers and regulating membrane biophysical properties, particularly lipid rafts, important signaling platforms. Altered sphingolipid levels have been implicated in many cardiovascular diseases, including hypertension, atherosclerosis, and diabetes mellitus–related conditions; however, molecular mechanisms by which ceramides impact endothelial functions remain poorly understood. In this regard, we generated mice defective of endothelial sphingolipid de novo biosynthesis by deleting the Sptlc2 (long chain subunit 2 of serine palmitoyltransferase)—the first enzyme of the pathway. Our study demonstrated that endothelial sphingolipid de novo production is necessary to regulate (1) signal transduction in response to NO agonists and, mainly via ceramides, (2) resting eNOS (endothelial NO synthase) phosphorylation, and (3) blood pressure homeostasis. Specifically, our findings suggest a prevailing role of C16:0-Cer in preserving vasodilation induced by tyrosine kinase and GPCRs (G-protein coupled receptors), except for Gq-coupled receptors, while C24:0- and C24:1-Cer control flow-induced vasodilation. Replenishing C16:0-Cer in vitro and in vivo reinstates endothelial cell signaling and vascular tone regulation. This study reveals an important role of locally produced ceramides, particularly C16:0-, C24:0-, and C24:1-Cer in vascular and blood pressure homeostasis, and establishes the endothelium as a key source of plasma ceramides. Clinically, specific plasma ceramides ratios are independent predictors of major cardiovascular events. Our data also suggest that plasma ceramides might be indicative of the diseased state of the endothelium.

Sphingolipids are both, fundamental structural components of the eukaryotic membranes and signaling molecules regulating a variety of biological functions. Highly bioactive lipids, ceramide and sphingosine-1-phosphate have emerged as important regulators of cardiovascular functions in health and disease. In this review we discuss recent insights into the role of sphingolipids, particularly ceramide and sphingosine-1-phosphate, in the pathophysiology of the cardiovascular system. We also highlight advances into the molecular mechanisms regulating serine palmitoyltransferase, the first and rate-limiting enzyme of de novo biosynthesis, with an emphasis on recently discovered inhibitors of serine palmitoyltransferase, ORMDL and NOGO-B proteins. Understanding the molecular mechanisms regulating this biosynthetic pathway may lead to the development of novel therapeutic approaches for the treatment of cardiovascular diseases.

Background: CD4 and Tetherin are stabilized through intrachain and interchain disulfide bonds, respectively. Results: CD4 and Tetherin retro-translocate from ER to cytosol with oxidized disulfide bridges as folded and multimeric molecules. Conclusion: Cysteines reduction is not a prerequisite for ER to cytosol dislocation. Significance: Our observations challenge the requirements of reduction and unfolding before dislocation.