Biology subjects

Parlato, R.

Publications and source records attributed to Parlato, R..

6 recordsLinked to original sources

bioRxiv2025

Probing How Anti-Huntingtin Antibodies Bind The Fibrillar Fuzzy Coat Using Solid-State NMR

Antibodies are critical for the immune response and serve as important tools due to their ability to recognize specific amino acid sequences, or epitopes. Based on the latter, they are utilized as diagnostic tools in biological and biomedical research. Huntingtons disease (HD) is a neurodegenerative condition caused by CAG repeat expansions in the huntingtin (HTT) gene and characterized by amyloid-like protein deposits in patients. Multiple anti-HTT antibodies are used in HD research for their ability to recognize specific HTT inclusions in both post-mortem tissue and in laboratory conditions. Some of the antibodies are seen as detectors of distinct structural motifs. However, most knowledge of their binding mechanism stems from studies of soluble monomers or short fragments of the epitopes, rather than the aggregated, misfolded target protein. Here, we investigate how MW8 antibodies interact with HTT exon 1 (HTTex1) fibrils, using solid-state NMR, electron microscopy, and complementary techniques. Magic angle spinning NMR revealed localized impacts of the antibody on exposed parts of the HTTex1 fibrils: the flanking segments that form its fuzzy coat. Antibody binding affected the structure and dynamics of the fuzzy coat, but also modulated the propensity for forming supramolecular fibril clusters, which has important implications for (reducing) cytotoxicity.

biochemistry↗

bioRxiv2025

Somatic mutations in ALS genes in the motor cortex of sporadic ALS patients

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration of cortical and spinal motor neurons. Mendelian germline mutations often cause familial ALS (fALS) but only approximately ten percent of sporadic ALS cases (sALS). We leveraged DNA and single cell RNA-sequencing data from autopsy tissue to explore the presence of somatic mosaic variants in sALS cases. Deep targeted panel sequencing of known ALS disease genes in motor cortex tissue revealed an enrichment of low allele frequency variants in sALS, but not in fALS with an identified monogenic cause. In silico analysis predicted increased pathogenicity of mosaic mutations in various known ALS mutational hot spots. In particular, we identified the somatic FUS variant p.E516X, located in an established hot spot for germline ALS mutations, which leads to nucleo-cytoplasmic mislocalization and aggregation typical for ALS FUS pathology. Additionally, we performed somatic variant calling on single cell RNA-sequencing data from sALS tissue and revealed a specific accumulation of somatic variants in excitatory neurons, reinforcing a neuron-autonomous disease initiation. Collectively, this study indicates that somatic mutations within the motor cortex, especially in excitatory neurons, may contribute to sALS development.

genetics↗

bioRxiv2024

Identifying similar populations across independent single cell studies without data integration

Supervised and unsupervised methods have emerged to address the complexity of single cell data analysis in the context of large pools of independent studies. Here, we present ClusterFoldSimilarity (CFS), a novel statistical method design to quantify the similarity between cell groups acroos any number of independent datasets, without the need for data correction or integration. By bypassing these processes, CFS avoids the introduction of artifacts and loss of information, offering a simple, efficient, and scalable solution. This method match groups of cells that exhibit conserved phenotypes across datasets, including different tissues and species, and in a multimodal scenario, including single-cell RNA-Seq, ATAC-Seq, single-cell proteomics, or, more broadly, data exhibiting differential abundance effects among groups of cells. Additionally, CFS performs feature selection, obtaining cross-dataset markers of the similar phenotypes observed, providing an inherent interpretability of relationships between cell populations. To showcase the effectiveness of our methodology we generated single-nuclei RNA-Seq data from the motor cortex and spinal cord of adult mice. By using CFS, we identified three distinct sub-populations of astrocytes conserved on both tissues. CFS includes various visualization methods for the interpretation of the similarity scores and similar cell populations.

bioinformatics↗

bioRxiv2022

Deep learning based image analysis identifies a DAT-negative subpopulation of dopaminergic neurons in the lateral Substantia nigra.

Here we present a deep learning-based image analysis platform (DLAP), tailored to autonomously quantify cell numbers, and fluorescence signals within cellular compartments, derived from RNAscope or immunohistochemistry. We utilized DLAP to analyse subtypes of tyrosine hydroxylase (TH)-positive dopaminergic midbrain neurons in mouse and human brain sections. These neurons modulate complex behaviour, and are differentially affected in Parkinsons and other diseases. DLAP allows the analysis of large cell numbers, and facilitates the identification of small cellular subpopulations. Specifically, we identified a small subpopulation of TH-positive neurons ([~]5%), mainly located in the very lateral Substantia nigra (SN), that was immunofluorescence-negative for the plasmalemma dopamine transporter (DAT), with [~]40% smaller cell bodies. These neurons were negative for aldehyde dehydrogenase 1A1, with a lower co-expression rate for dopamine-D2-autoreceptors, but a [~]7-fold higher likelihood of calbindin-d28k co-expression ([~]70%). Our results have important implications, as DAT is crucial for dopamine-signalling, and is commonly used as a marker for dopaminergic SN neurons.

neuroscience↗

bioRxiv2021

Nucleolar stress controls mutant Huntingtin toxicity and monitors Huntington disease progression

Transcriptional and cellular stress surveillance deficits are hallmarks of Huntingtons disease (HD), a fatal autosomal dominant neurodegenerative disorder, caused by a pathological expansion of CAG repeats in the Huntingtin (HTT) gene. The nucleolus, a dynamic nuclear biomolecular condensate and the site of ribosomal RNA (rRNA) transcription, is implicated in the cellular stress response and in protein quality control. While the exact pathomechanisms of HD remain unclear, the impact of nucleolar dysfunction on HD pathophysiology in vivo is elusive. Here we identified aberrant maturation of rRNA and decreased translational rate in association with human mutant Huntingtin (mHTT) expression. Genetic disruption of nucleolar integrity in vulnerable striatal neurons of the R6/2 HD mouse model decreases mHTT disperse state in the nucleus, exacerbating the motor deficits. The protein nucleophosmin 1 (NPM1), important for nucleolar integrity and rRNA maturation, loses its nucleolar localization. NPM1 de-localization occurs in the striatum and in the skeletal muscle of the progressive zQ175 knock-in HD mouse model, mimicking the phenotype of HD patients in skeletal muscle biopsies. Taken together, we showed that nucleolar integrity regulates the formation of mHTT inclusions in vivo, and identified NPM1 as a novel, readily detectable peripheral histopathological marker of HD progression.

neuroscience↗

bioRxiv2019

Structural fuzziness of the RNA-organizing protein SERF1a determines a toxic gain-of-interaction

Members of the MOAG/SERF protein family have been attributed a neuropathologic significance because of their ability to enhance the proteotoxic polymerization of amyloidogenic proteins such as alpha-Synuclein (aSyn). However, the cellular function remains unknown. Here, we identify SERF1a as an RNA-organizing protein i) with no structural homology to canonical RNA-binding proteins, ii) with an RNA-chaperoning activity which favours the incorporation of RNA into nucleoli and liquid-like artificial RNA-organelles, and iii) with a high degree of conformational disorder in the RNA-bound state. We demonstrate that this type of structural fuzziness determines an undifferentiated interaction of SERF1a with aSyn and RNA. Both molecules bind to one identical, positively charged site of SERF1a by an analogous electrostatic binding mode, with similar binding affinities, and without any observable disorder-to-order transition. The absence of primary or secondary structure discriminants results in SERF1a being unable to distinguish between nucleic acid and amyloidogenic protein, leading the pro-amyloid aSyn:SERF1a interaction to prevail under conditions of cellular stress. We suggest that structural fuzziness of SERF1a accounts for an adverse gain-of-interaction which favours toxic binding to aSyn at the expense of non-toxic RNA binding, therefore promoting a functionally distorted and pathogenic process. Our results provide a direct link between structural disorder, amyloid aggregation and the malfunctioning of RNA-associated cellular processes, three hallmarks of neurodegenerative diseases.

biochemistry↗