The neuronal microtubule-associated protein tau undergoes multiple post-translational modifications, which dynamically modulate its molecular functions and biochemical features in space and time. Among them, we have recently reported that a conserved lysine residue mapping to the microtubule-binding domain of the protein (K306 in mouse and K317 in human) is differentially methylated in a model of chronic autoimmune demyelination. In contrast with other well-studied tau post-translational modifications such as phosphorylation, lysine methylation is far less investigated and its specific impact on tau biology is not fully understood. Here we performed a comprehensive analysis of the effects of K317 methylation on key tau features. By combining in silico simulations with in vitro biochemical assays and live-cell imaging, we show that methylated tau is more prone to self-assembly into insoluble structures. Moreover, we demonstrate that K317 methylation affects the stabilization activity of tau on microtubule dynamics. Lastly, we highlight a role for K317 methylation in regulating both neuronal differentiation and cell proliferation. Altogether, these findings shed light on the biology of an overlooked tau post-translational modification as well as on the fine tuning of tau functionality in health and disease.

The human leukocyte antigen (HLA) locus encodes a large group of proteins governing adaptive and innate immune responses. Among them, HLA class II proteins form α/β heterodimers on the membrane of professional antigen-presenting cells (APCs), where they display both, self and pathogen-derived exogenous antigens to CD4⁺ T lymphocytes. We have previously shown that a shorter HLA-DRA isoform (sHLA-DRA) lacking 25 amino acids can be presented onto the cell membrane via binding to canonical HLA-DR2 heterodimers. Here, we employed atomistic molecular dynamics simulations to decipher the binding position of sHLA-DRA and its structural impact on functional regions of the HLA-DR2 molecule. We show that a loop region exposed only in the short isoform (residues R69 to G83) is responsible for binding to the outer domain of the HLA-DR2 peptide-binding site, and experimentally validated the critical role of F76 in mediating such interaction. Additionally, sHLA-DRA allosterically modifies the peptide-binding pocket conformation. In summary, this study unravels key molecular mechanisms underlying sHLA-DRA function, providing important insights into the role of full-length proteins in structural modulation of HLA class II receptors.

Background

MicroRNAs (miRNAs) belong to a class of evolutionary conserved, non-coding small RNAs with regulatory functions on gene expression. They negatively affect the expression of target genes by promoting either RNA degradation or translational inhibition. In recent years, converging studies have identified miRNAs as key regulators of oligodendrocyte (OL) functions. OLs are the cells responsible for the formation and maintenance of myelin in the central nervous system (CNS) and represent a principal target of the autoimmune injury in multiple sclerosis (MS).

Methods

MiRAP is a novel cell-specific miRNA affinity-purification technique which relies on genetically tagging Argonaut 2 (AGO2), an enzyme involved in miRNA processing. Here, we exploited miRAP potentiality to characterize OL-specific miRNA dynamics in the MS model experimental autoimmune encephalomyelitis (EAE).

Results

We show that 20 miRNAs are differentially regulated in OLs upon transition from pre-symptomatic EAE stages to disease peak. Subsequent in vitro differentiation experiments demonstrated that a sub-group of them affects the OL maturation process, mediating either protective or detrimental signals. Lastly, transcriptome profiling highlighted the endocytosis, ferroptosis, and FoxO cascades as the pathways associated with miRNAs mediating or inhibiting OL maturation.

Conclusions

Altogether, our work supports a dual role for miRNAs in autoimmune demyelination. In particular, the enrichment in miRNAs mediating pro-myelinating signals suggests an active involvement of these non-coding RNAs in the homeostatic response toward neuroinflammatory injury.

Animal models are widely employed in basic research to test mechanistic hypotheses in a complex biological environment as well as to evaluate the therapeutic potential of candidate compounds in preclinical settings. Rodents, and in particular mice, represent the most common in vivo models for their small size, short lifespan and possibility to manipulate their genome. Over time, a typical laboratory will develop a substantial number of inbred strains and transgenic mouse lines, requiring a substantial effort, in both logistic and economic terms, to maintain an animal colony for research purposes and to safeguard the integrity of results. To meet this need, here we present TopoDB, a robust and extensible web-based platform for the rational management of laboratory animals. TopoDB allows an easy tracking of individual animals within the colony and breeding protocols as well as the convenient storage of both genetic and phenotypic data generated in the different experiments. Altogether, these features facilitate and enhance the design of in vivo research, thus reducing the number of necessary animals and the housing costs. In summary, TopoDB represents a novel valuable tool in modern biomedical research. Database URL: https://github.com/UCSF-MS-DCC/TopoDB.

Ataxin-1 (ATXN1) is a ubiquitous polyglutamine protein expressed primarily in the nucleus where it binds chromatin and functions as a transcriptional repressor. Mutant forms of ataxin-1 containing expanded glutamine stretches cause the movement disorder spinocerebellar ataxia type 1 (SCA1) through a toxic gain-of-function mechanism in the cerebellum. Conversely, ATXN1 loss-of-function is implicated in cancer development and Alzheimer's disease (AD) pathogenesis. ATXN1 was recently nominated as a susceptibility locus for multiple sclerosis (MS). Here, we show that Atxn1-null mice develop a more severe experimental autoimmune encephalomyelitis (EAE) course compared to wildtype mice. The aggravated phenotype is mediated by increased T helper type 1 (Th1) cell polarization, which in turn results from the dysregulation of B cell activity. Ataxin-1 ablation in B cells leads to aberrant expression of key costimulatory molecules involved in proinflammatory T cell differentiation, including cluster of differentiation (CD)44 and CD80. In addition, comprehensive phosphoflow cytometry and transcriptional profiling link the exaggerated proliferation of ataxin-1 deficient B cells to the activation of extracellular signal-regulated kinase (ERK) and signal transducer and activator of transcription (STAT) pathways. Lastly, selective deletion of the physiological binding partner capicua (CIC) demonstrates the importance of ATXN1 native interactions for correct B cell functioning. Altogether, we report a immunomodulatory role for ataxin-1 and provide a functional description of the ATXN1 locus genetic association with MS risk.