Ral GTPases are critical regulators of spinal cord myelination and homeostasis.

Efficient myelination supports nerve conduction and axonal health throughout life. In the central nervous system, oligodendrocytes (OLs) carry out this demanding anabolic duty in part through biosynthetic pathways controlled by mTOR. We identify Ral GTPases as critical regulators of mouse spinal cord myelination and myelin maintenance. Ablation of Ral GTPases (RalA, RalB) in OL-lineage cells impairs timely onset and radial growth of developmental myelination, accompanied by increased endosomal/lysosomal abundance. Further examinations, including transcriptomic analyses of Ral-deficient OLs, were consistent with mTORC1-related deficits. However, deletion of the mTOR signaling-repressor Pten in Ral-deficient OL-lineage cells is unable to rescue mTORC1 activation or developmental myelination deficiencies. Induced deletion of Ral GTPases in OLs of adult mice results in late-onset myelination defects and tissue degeneration. Together, our data indicate critical roles for Ral GTPases to promote developmental spinal cord myelination, to ensure accurate mTORC1 signaling, and to protect the healthy state of myelin-axon units over time.

Transcriptional profiling of mouse peripheral nerves to the single-cell level to build a sciatic nerve ATlas (SNAT).

Peripheral nerves are organ-like structures containing diverse cell types to optimize function. This interactive assembly includes mostly axon-associated Schwann cells, but also endothelial cells of supporting blood vessels, immune system-associated cells, barrier-forming cells of the perineurium surrounding and protecting nerve fascicles, and connective tissue-resident cells within the intra-fascicular endoneurium and inter-fascicular epineurium. We have established transcriptional profiles of mouse sciatic nerve-inhabitant cells to foster the fundamental understanding of peripheral nerves. To achieve this goal, we have combined bulk RNA sequencing of developing sciatic nerves up to the adult with focused bulk and single-cell RNA sequencing of Schwann cells throughout postnatal development, extended by single-cell transcriptome analysis of the full sciatic nerve both perinatally and in the adult. The results were merged in the transcriptome resource Sciatic Nerve ATlas (SNAT: https://www.snat.ethz.ch). We anticipate that insights gained from our multi-layered analysis will serve as valuable interactive reference point to guide future studies.

Mice carrying an analogous heterozygous dynamin 2 K562E mutation that causes neuropathy in humans develop predominant characteristics of a primary myopathy.

Some mutations affecting dynamin 2 (DNM2) can cause dominantly inherited Charcot-Marie-Tooth (CMT) neuropathy. Here, we describe the analysis of mice carrying the DNM2 K562E mutation which has been associated with dominant-intermediate CMT type B (CMTDIB). Contrary to our expectations, heterozygous DNM2 K562E mutant mice did not develop definitive signs of an axonal or demyelinating neuropathy. Rather, we found a primary myopathy-like phenotype in these mice. A likely interpretation of these results is that the lack of a neuropathy in this mouse model has allowed the unmasking of a primary myopathy due to the DNM2 K562E mutation which might be overshadowed by the neuropathy in humans. Consequently, we hypothesize that a primary myopathy may also contribute to the disease mechanism in some CMTDIB patients. We propose that these findings should be considered in the evaluation of patients, the determination of the underlying disease processes and the development of tailored potential treatment strategies.

Ral GTPases in Schwann cells promote radial axonal sorting in the peripheral nervous system.

Small GTPases of the Rho and Ras families are important regulators of Schwann cell biology. The Ras-like GTPases RalA and RalB act downstream of Ras in malignant peripheral nerve sheath tumors. However, the physiological role of Ral proteins in Schwann cell development is unknown. Using transgenic mice with ablation of one or both Ral genes, we report that Ral GTPases are crucial for axonal radial sorting. While lack of only one Ral GTPase was dispensable for early peripheral nerve development, ablation of both RalA and RalB resulted in persistent radial sorting defects, associated with hallmarks of deficits in Schwann cell process formation and maintenance. In agreement, ex vivo-cultured Ral-deficient Schwann cells were impaired in process extension and the formation of lamellipodia. Our data indicate further that RalA contributes to Schwann cell process extensions through the exocyst complex, a known effector of Ral GTPases, consistent with an exocyst-mediated function of Ral GTPases in Schwann cells.

CNS myelination and remyelination depend on fatty acid synthesis by oligodendrocytes.

Oligodendrocytes (OLs) support neurons and signal transmission in the central nervous system (CNS) by enwrapping axons with myelin, a lipid-rich membrane structure. We addressed the significance of fatty acid (FA) synthesis in OLs by depleting FA synthase (FASN) from OL progenitor cells (OPCs) in transgenic mice. While we detected no effects in proliferation and differentiation along the postnatal OL lineage, we found that FASN is essential for accurate myelination, including myelin growth. Increasing dietary lipid intake could partially compensate for the FASN deficiency. Furthermore, FASN contributes to correct myelin lipid composition and stability of myelinated axons. Moreover, we depleted FASN specifically in adult OPCs to examine its relevance for remyelination. Applying lysolecithin-induced focal demyelinating spinal cord lesions, we found that FA synthesis is essential to sustain adult OPC-derived OLs and efficient remyelination. We conclude that FA synthesis in OLs plays key roles in CNS myelination and remyelination.

Schwann cells, but not Oligodendrocytes, Depend Strictly on Dynamin 2 Function.

Myelination requires extensive plasma membrane rearrangements, implying that molecules controlling membrane dynamics play prominent roles. The large GTPase dynamin 2 (DNM2) is a well-known regulator of membrane remodeling, membrane fission, and vesicular trafficking. Here, we genetically ablated Dnm2 in Schwann cells (SCs) and in oligodendrocytes of mice. Dnm2 deletion in developing SCs resulted in severely impaired axonal sorting and myelination onset. Induced Dnm2 deletion in adult SCs caused a rapidly-developing peripheral neuropathy with abundant demyelination. In both experimental settings, mutant SCs underwent prominent cell death, at least partially due to cytokinesis failure. Strikingly, when Dnm2 was deleted in adult SCs, non-recombined SCs still expressing DNM2 were able to remyelinate fast and efficiently, accompanied by neuropathy remission. These findings reveal a remarkable self-healing capability of peripheral nerves that are affected by SC loss. In the central nervous system, however, we found no major defects upon Dnm2 deletion in oligodendrocytes.