Lipid-anchored proteasomes control membrane protein homeostasis.

Protein degradation in eukaryotic cells is mainly carried out by the 26S proteasome, a macromolecular complex not only present in the cytosol and nucleus but also associated with various membranes. How proteasomes are anchored to the membrane and the biological meaning thereof have been largely unknown in higher organisms. Here, we show that N-myristoylation of the Rpt2 subunit is a general mechanism for proteasome-membrane interaction. Loss of this modification in the Rpt2-G2A mutant cells leads to profound changes in the membrane-associated proteome, perturbs the endomembrane system, and undermines critical cellular processes such as cell adhesion, endoplasmic reticulum-associated degradation and membrane protein trafficking. Rpt2G2A/G2A homozygous mutation is embryonic lethal in mice and is sufficient to abolish tumor growth in a nude mice xenograft model. These findings have defined an evolutionarily conserved mechanism for maintaining membrane protein homeostasis and underscored the significance of compartmentalized protein degradation by myristoyl-anchored proteasomes in health and disease.

GDF11 slows excitatory neuronal senescence and brain ageing by repressing p21.

As a major neuron type in the brain, the excitatory neuron (EN) regulates the lifespan in C. elegans. How the EN acquires senescence, however, is unknown. Here, we show that growth differentiation factor 11 (GDF11) is predominantly expressed in the EN in the adult mouse, marmoset and human brain. In mice, selective knock-out of GDF11 in the post-mitotic EN shapes the brain ageing-related transcriptional profile, induces EN senescence and hyperexcitability, prunes their dendrites, impedes their synaptic input, impairs object recognition memory and shortens the lifespan, establishing a functional link between GDF11, brain ageing and cognition. In vitro GDF11 deletion causes cellular senescence in Neuro-2a cells. Mechanistically, GDF11 deletion induces neuronal senescence via Smad2-induced transcription of the pro-senescence factor p21. This work indicates that endogenous GDF11 acts as a brake on EN senescence and brain ageing.

Lipid-anchored Proteasomes Control Membrane Protein Homeostasis.

Protein degradation in eukaryotic cells is mainly carried out by the 26S proteasome, a macromolecular complex not only present in the cytosol and nucleus but also associated with various membranes. How proteasomes are anchored to the membrane and the biological meaning thereof have been largely unknown in higher organisms. Here we show that N-myristoylation of the Rpt2 subunit is a general mechanism for proteasome-membrane interaction. Loss of this modification in the Rpt2-G2A mutant cells leads to profound changes in the membrane-associated proteome, perturbs the endomembrane system and undermines critical cellular processes such as cell adhesion, endoplasmic reticulum-associated degradation (ERAD) and membrane protein trafficking. Rpt2 G2A/G2A homozygous mutation is embryonic lethal in mice and is sufficient to abolish tumor growth in a nude mice xenograft model. These findings have defined an evolutionarily conserved mechanism for maintaining membrane protein homeostasis and underscored the significance of compartmentalized protein degradation by m yristoyl- a nchored p roteasomes (MAPs) in health and disease.

A novel NF2 splicing mutant causes neurofibromatosis type 2 via liquid-liquid phase separation with large tumor suppressor and Hippo pathway.

Neurofibromatosis type 2 is an autosomal dominant multiple neoplasia syndrome and is usually caused by mutations in the neurofibromin 2 (NF2) gene, which encodes a tumor suppressor and initiates the Hippo pathway. However, the mechanism by which NF2 functions in the Hippo pathway isn't fully understood. Here we identified a NF2 c.770-784del mutation from a neurofibromatosis type 2 family. MD simulations showed that this mutation significantly changed the structure of the F3 module of the NF2-FERM domain. Functional assays indicated that the NF2 c.770-784del variant formed LLPS in the cytoplasm with LATS to restrain LATS plasma membrane localization and inactivated the Hippo pathway. Besides, this deletion partly caused a skipping of exon 8 and reduced the protein level of NF2, collectively promoting proliferation and tumorigenesis of meningeal cells. We identified an unrecognized mechanism of LLPS and splicing skipping for the NF2-induced Hippo pathway, which provided new insight into the pathogenesis of neurofibromatosis type 2.

Inhibition of RIPK1 by ZJU-37 promotes oligodendrocyte progenitor proliferation and remyelination via NF-κB pathway.

Receptor interacting serine/threonine protein kinase 1 (RIPK1) activation and necroptosis have been genetically and mechanistically linked with human multiple sclerosis and neurodegenerative diseases for which demyelination is a common key pathology. Demyelination can be healed through remyelination which is mediated by new oligodendrocytes derived from the adult oligodendrocyte progenitor cells (OPCs). Unfortunately, the efficiency of remyelination declines with progressive aging partially due to the depletion of OPCs following chronic or repeated demyelination. However, to our knowledge, so far there is no drug which enhances proliferation of OPCs, and it is unknown whether inhibiting RIPK1 activity directly affect OPCs, the central player of remyelination. Using TNFα induced RIPK1-dependent necroptosis in Jurkat FADD-/- cells as a cell death assay, we screened from 2112 FDA-approved drugs and the drug candidates of new RIPK1 inhibitors selected by ourselves, and identified ZJU-37, a small molecule modified by introducing an amide bond to Nec-1s, is a new RIPK1 kinase inhibitor with higher potency than Nec-1s which has the best reported potency. We unveil in addition to protecting myelin from demyelination and axons from degeneration, ZJU-37 exhibits a new role on promoting proliferation of OPCs and enhancing remyelination by inhibiting RIPK1 kinase activity with higher potency than Nec-1s. Mechanistically, ZJU-37 promotes proliferation of OPCs by enhancing the transcription of platelet derived growth factor receptor alpha via NF-κB pathway. This work identifies ZJU-37 as a new drug candidate which enhances remyelination by promoting proliferation of OPCs, paving the way for a potential drug to enhance myelin repair.

Impaired metabolism of oligodendrocyte progenitor cells and axons in demyelinated lesion and in the aged CNS.

The key pathology of multiple sclerosis (MS) comprises demyelination, axonal damage, and neuronal loss, and when MS develops into the progressive phase it is essentially untreatable. Identifying new targets in both axons and oligodendrocyte progenitor cells (OPCs) and rejuvenating the aged OPCs holds promise for this unmet medical need. We summarize here the recent evidence showing that mitochondria in both axons and OPCs are impaired, and lipid metabolism of OPCs within demyelinated lesion and in the aged CNS is disturbed. Given that emerging evidence shows that rewiring cellular metabolism regulates stem cell aging, to protect axons from degeneration and promote differentiation of OPCs, we propose that restoring the impaired metabolism of both OPCs and axons in the aged CNS in a synergistic way could be a promising strategy to enhance remyelination in the aged CNS, leading to novel drug-based approaches to treat the progressive phase of MS.

Restoring nuclear entry of Sirtuin 2 in oligodendrocyte progenitor cells promotes remyelination during ageing.

The age-dependent decline in remyelination potential of the central nervous system during ageing is associated with a declined differentiation capacity of oligodendrocyte progenitor cells (OPCs). The molecular players that can enhance OPC differentiation or rejuvenate OPCs are unclear. Here we show that, in mouse OPCs, nuclear entry of SIRT2 is impaired and NAD+ levels are reduced during ageing. When we supplement ß-nicotinamide mononucleotide (ß-NMN), an NAD+ precursor, nuclear entry of SIRT2 in OPCs, OPC differentiation, and remyelination were rescued in aged animals. We show that the effects on myelination are mediated via the NAD+-SIRT2-H3K18Ac-ID4 axis, and SIRT2 is required for rejuvenating OPCs. Our results show that SIRT2 and NAD+ levels rescue the aged OPC differentiation potential to levels comparable to young age, providing potential targets to enhance remyelination during ageing.

Neutralization of Hv1/HVCN1 With Antibody Enhances Microglia/Macrophages Myelin Clearance by Promoting Their Migration in the Brain.

Microglia dynamically monitor the microenvironment of the central nervous system (CNS) by constantly extending and retracting their processes in physiological conditions, and microglia/macrophages rapidly migrate into lesion sites in response to injuries or diseases in the CNS. Consequently, their migration ability is fundamentally important for their proper functioning. However, the mechanisms underlying their migration have not been fully understood. We wonder whether the voltage-gated proton channel HVCN1 in microglia/macrophages in the brain plays a role in their migration. We show in this study that in physiological conditions, microglia and bone marrow derived macrophage (BMDM) express HVCN1 with the highest level among glial cells, and upregulation of HVCN1 in microglia/macrophages is presented in multiple injuries and diseases of the CNS, reflecting the overactivation of HVCN1. In parallel, myelin debris accumulation occurs in both the focal lesion and the site where neurodegeneration takes place. Importantly, both genetic deletion of the HVCN1 gene in cells in vitro and neutralization of HVCN1 with antibody in the brain in vivo promotes migration of microglia/macrophages. Furthermore, neutralization of HVCN1 with antibody in the brain in vivo promotes myelin debris clearance by microglia/macrophages. This study uncovers a new role of HVCN1 in microglia/macrophages, coupling the proton channel HVCN1 to the migration of microglia/macrophages for the first time.

Loss of Growth Differentiation Factor 11 Shortens Telomere Length by Downregulating Telomerase Activity.

Maintenance of telomere length is essential to delay replicative cellular senescence. It is controversial on whether growth differentiation factor 11 (GDF11) can reverse cellular senescence, and this work aims to establish the causality between GDF11 and the telomere maintenance unequivocally. Using CRISPR/Cas9 technique and a long-term in vitro culture model of cellular senescence, we show here that in vitro genetic deletion of GDF11 causes shortening of telomere length, downregulation of telomeric reverse transcriptase (TERT) and telomeric RNA component (TERC), the key enzyme and the RNA component for extension of the telomere, and reduction of telomerase activity. In contrast, both recombinant and overexpressed GDF11 restore the transcription of TERT in GDF11KO cells to the wild-type level. Furthermore, loss of GDF11-induced telomere shortening is likely caused by enhancing the nuclear entry of SMAD2 which inhibits the transcription of TERT and TERC. Our results provide the first proof-of-cause-and-effect evidence that endogenous GDF11 plays a causal role for proliferative cells to maintain telomere length, paving the way for potential rejuvenation of the proliferative cells, tissues, and organs.