Ganglioside lipidomics of CNS myelination using direct infusion shotgun mass spectrometry.

Gangliosides are present and concentrated in axons and implicated in axon-myelin interactions, but how ganglioside composition changes during myelin formation is not known. Here, we present a direct infusion (shotgun) lipidomics method to analyze gangliosides in small amounts of tissue reproducibly and with high sensitivity. We resolve the mouse ganglioside lipidome during development and adulthood and determine the ganglioside content of mice lacking the St3gal5 and B4galnt1 genes that synthesize most ganglioside species. Our results reveal substantial changes in the ganglioside lipidome during the formation of myelinated nerve fibers. In sum, we provide insights into the CNS ganglioside lipidome with a quantitative and sensitive mass spectrometry method. Since this method is compatible with global lipidomic profiling, it will provide insights into ganglioside function in physiology and pathology.

The pseudoprotease iRhom1 controls ectodomain shedding of membrane proteins in the nervous system.

Proteolytic ectodomain shedding of membrane proteins is a fundamental mechanism to control the communication between cells and their environment. A key protease for membrane protein shedding is ADAM17, which requires a non-proteolytic subunit, either inactive Rhomboid 1 (iRhom1) or iRhom2 for its activity. While iRhom1 and iRhom2 are co-expressed in most tissues and appear to have largely redundant functions, the brain is an organ with predominant expression of iRhom1. Yet, little is known about the spatio-temporal expression of iRhom1 in mammalian brain and about its function in controlling membrane protein shedding in the nervous system. Here, we demonstrate that iRhom1 is expressed in mouse brain from the prenatal stage to adulthood with a peak in early postnatal development. In the adult mouse brain iRhom1 was widely expressed, including in cortex, hippocampus, olfactory bulb, and cerebellum. Proteomic analysis of the secretome of primary neurons using the hiSPECS method and of cerebrospinal fluid, obtained from iRhom1-deficient and control mice, identified several membrane proteins that require iRhom1 for their shedding in vitro or in vivo. One of these proteins was 'multiple-EGF-like-domains protein 10' (MEGF10), a phagocytic receptor in the brain that is linked to the removal of amyloid ß and apoptotic neurons. MEGF10 was further validated as an ADAM17 substrate using ADAM17-deficient mouse embryonic fibroblasts. Taken together, this study discovers a role for iRhom1 in controlling membrane protein shedding in the mouse brain, establishes MEGF10 as an iRhom1-dependent ADAM17 substrate and demonstrates that iRhom1 is widely expressed in murine brain.

Two adhesive systems cooperatively regulate axon ensheathment and myelin growth in the CNS.

Central nervous system myelin is a multilayered membrane produced by oligodendrocytes to increase neural processing speed and efficiency, but the molecular mechanisms underlying axonal selection and myelin wrapping are unknown. Here, using combined morphological and molecular analyses in mice and zebrafish, we show that adhesion molecules of the paranodal and the internodal segment work synergistically using overlapping functions to regulate axonal interaction and myelin wrapping. In the absence of these adhesive systems, axonal recognition by myelin is impaired with myelin growing on top of previously myelinated fibers, around neuronal cell bodies and above nodes of Ranvier. In addition, myelin wrapping is disturbed with the leading edge moving away from the axon and in between previously formed layers. These data show how two adhesive systems function together to guide axonal ensheathment and myelin wrapping, and provide a mechanistic understanding of how the spatial organization of myelin is achieved.