Biology subjects

Gillis, T.

Publications and source records attributed to Gillis, T..

12 recordsLinked to original sources

bioRxiv2026

Somatic CRISPR editing of Msh3 mitigates Huntington's disease pathology in mice

Huntingtons disease (HD) is a fatal, dominantly inherited neurodegenerative disorder caused by a CAG repeat expansion in Huntingtin (HTT) exon 1. Further progressive CAG repeat expansion occurs in somatic cells, particularly in neurons, and drives the timing of clinical onset. Therefore, therapeutic strategies to slow somatic expansion are predicted to be disease-modifying. Somatic CAG expansion is driven by mismatch repair protein MSH3, a leading therapeutic target supported by human genetic data. To gain insight into the impact of targeting MSH3 at different stages of the disease process we used somatic CRISPR-Cas9 editing to knock out Msh3 in HttQ111 mice at ages of 6, 16, 24 weeks exhibiting progressively increasing somatic expansion. Intervention at all three ages slowed striatal CAG expansion, reduced nuclear huntingtin pathology and suppressed transcriptional dysregulation, with earlier intervention having greater impact. Msh3 knockout also suppressed the production of the exon 1 Htt1a transcript. The results of our study provide important preclinical information relevant to an MSH3 therapeutic in humans that would be expected to impact a subset of cells in the brain, provide insight into the influence of timing of intervention on therapeutic effectiveness and deepen our understanding of how targeting MSH3 could alter the trajectory of HD.

genetics↗

bioRxiv2026

TRACE: Open-Source Software for Quantifying Somatic Variation of Tandem Repeats by Capillary Electrophoresis

Expanded short tandem DNA repeats are implicated in over 60 human disorders. In many, somatic instability (SI) of the repeat plays a critical role in disease pathogenesis. For example, SI in vulnerable neurons is a key driver of clinical symptoms in Huntingtons disease. Quantifying SI has traditionally relied on PCR followed by capillary electrophoresis, with metrics describing the shape of repeat size distributions, such as the expansion index. However, current tools often require costly proprietary software, are time-consuming, and rely on custom pipelines that vary between labs. To address these challenges, we developed Tandem Repeats Analysis by Capillary Electrophoresis (TRACE), an open-source software that processes fragment analysis data end-to-end, from raw files to SI metrics. Additionally, we created an associated web app TRACE-shiny (https://traceshiny.mgh.harvard.edu/), for interactive usage. Outputs from TRACE benchmarked against published datasets confirm its utility for studying genetic and pharmacological modifiers of SI. TRACE eliminates the need for proprietary software or custom pipelines, making advanced tools for analysis of somatic repeat expansion widely accessible.

genetics↗

bioRxiv2025

Huntington's disease LIG1 modifier variant increases ligase fidelity and suppresses somatic CAG repeat expansion

Huntingtons disease (HD) is a fatal neurodegenerative disorder caused by inheriting an expanded CAG repeat tract in the huntingtin gene (HTT) that further expands in somatic cells over an individuals lifetime. Genome-wide association studies have provided critical insight into factors that modify the course of disease. These include DNA repair genes that alter the rate of somatic expansion and other genes that do not appear to directly influence this process. One modifier gene is DNA ligase 1 (LIG1), in which a variant specifying a lysine to asparagine substitution (K845N) is associated with a profound (7-8 year) delay in the onset of motor signs. Here, we have taken a multifaceted approach to gain insight into the protective nature of this variant in HD. We demonstrate using in vitro ligase assays and enzyme kinetics that K845N enhances discrimination towards mismatched substrates and increases repair fidelity. Consistent with increased ligation fidelity, K845N confers protection against oxidative stress in cell-based assays. Finally, we demonstrate that the mouse LIG1 K843N orthologue suppresses somatic CAG expansion in HD knock-in mice. Overall, our data provide evidence that altered LIG1 function due to the K845N substitution may contribute to HD clinical delay by slowing somatic expansion in the brain and protecting the genome globally against damage. Significantly, our results provide a mechanistic foundation for considering DNA ligase fidelity as a therapeutic target in HD and potentially in other trinucleotide repeat disorders. Significance StatementWe analyzed a missense variant in DNA Ligase 1 (K845N) that is associated with a profound delay in the onset of Huntingtons disease (HD). We find that K845N enhances substrate discrimination towards mismatched substrates, thus increasing repair fidelity, conferring protection against oxidative stress and slows somatic expansion of the HD CAG repeat. Our observations provide insight into underlying mechanisms of disease modification and suggest avenues that can be harnessed for disease-modifying therapeutic intervention. Classification: Biological Sciences, Genetics

genetics↗

bioRxiv2025

MSH3 is a genetic modifier of somatic repeat instability in X-linked dystonia parkinsonism

X-linked dystonia parkinsonism (XDP) is a progressive adult-onset neurogenerative disorder caused by the insertion of a SINE-VNTR-Alu (SVA) retrotransposon in TAF1 gene. One element of the SVA is a tandem polymorphic CCCTCT repeat tract whose length inversely correlates with the age of disease onset. Previous observations that the repeat exhibits length-dependent somatic expansion and that XDP onset is modified by variation in DNA repair gene MSH3 indicated that somatic repeat expansion is an important disease driver. Here, we sought to uncover genetic modifiers of CCCTCT instability in XDP patients and to provide a mechanistic link between somatic instability and disease. We determined quantitative metrics of both repeat expansion and repeat contraction in blood. Using genetic association analyses of exome sequencing data, as well as directed sequencing of a variant MSH3 repeat, we found that MSH3 modifies repeat expansion and contraction in blood as well as age at onset. MSH3 alleles associated with earlier disease onset were associated with more expansion and less contraction. Conversely, alleles associated with later disease onset were associated with less expansion and more contraction. Notably, MSH3 repeat alleles were also similarly associated with expansion and contraction in brain tissues. Our findings provide key evidence that MSH3s role(s) in CCCTCT repeat dynamics underlies its impact on clinical disease and indicate that therapeutic strategies to lower or inhibit MSH3 are predicted to both slow CCCTCT expansion and promote CCCTCT contraction, impacting the disease course prior to clinical onset.

genetics↗

bioRxiv2024

Genetic modifiers of somatic expansion and clinical phenotypes in Huntington's disease reveal shared and tissue-specific effects

Huntingtons disease (HD), due to expansion of a CAG repeat in HTT, is representative of a growing number of disorders involving somatically unstable short tandem repeats. We find that overlapping and distinct genetic modifiers of clinical landmarks and somatic expansion in blood DNA reveal an underlying complexity and cell-type specificity to the mismatch repair-related processes that influence disease timing. Differential capture of non-DNA-repair gene modifiers by multiple measures of cognitive and motor dysfunction argues additionally for cell-type specificity of pathogenic processes. Beyond trans modifiers, differential effects are also illustrated at HTT by a 5-UTR variant that promotes somatic expansion in blood without influencing clinical HD, while, even after correcting for uninterrupted CAG length, a synonymous sequence change at the end of the CAG repeat dramatically hastens onset of motor signs without increasing somatic expansion. Our findings are directly relevant to therapeutic suppression of somatic expansion in HD and related disorders and provide a route to define the individual neuronal cell types that contribute to different HD clinical phenotypes.

genetics↗

bioRxiv2024

Identification of genetic modifiers of Huntington's disease somatic CAG repeat instability by in vivo CRISPR-Cas9 genome editing

Huntingtons disease (HD), one of >50 inherited repeat expansion disorders (Depienne and Mandel, 2021), is a dominantly-inherited neurodegenerative disease caused by a CAG expansion in HTT (The Huntingtons Disease Collaborative Research Group, 1993). Inherited CAG repeat length is the primary determinant of age of onset, with human genetic studies underscoring that the property driving disease is the CAG length-dependent propensity of the repeat to further expand in brain (Swami et al., 2009; GeM-HD, 2015; Hensman Moss et al., 2017; Ciosi et al., 2019; GeM-HD, 2019; Hong et al., 2021). Routes to slowing somatic CAG expansion therefore hold great promise for disease-modifying therapies. Several DNA repair genes, notably in the mismatch repair (MMR) pathway, modify somatic expansion in HD mouse models (Wheeler and Dion, 2021). To identify novel modifiers of somatic expansion, we have used CRISPR-Cas9 editing in HD knock-in mice to enable in vivo screening of expansion-modifier candidates at scale. This has included testing of HD onset modifier genes emerging from human genome-wide association studies (GWAS), as well as interactions between modifier genes, thereby providing new insight into pathways underlying CAG expansion and potential therapeutic targets.

genetics↗

bioRxiv2023

A parametrized two-domain thermodynamic model explains diverse mutational effects on protein allostery

New experimental findings continue to challenge our understanding of protein allostery. Recent deep mutational scanning study showed that allosteric hotspots in the tetracycline repressor (TetR) and its homologous transcriptional factors are broadly distributed rather than spanning well-defined structural pathways as often assumed. Moreover, hotspot mutation-induced allostery loss was rescued by distributed additional mutations in a degenerate fashion. Here, we develop a two-domain thermodynamic model for TetR, which readily rationalizes these intriguing observations. The model accurately captures the in vivo activities of various mutants with changes in physically transparent parameters, allowing the data-based quantification of mutational effects using statistical inference. Our analysis reveals the intrinsic connection of intra- and inter-domain properties for allosteric regulation and illustrate epistatic interactions that are consistent with structural features of the protein. The insights gained from this study into the nature of two-domain allostery are expected to have broader implications for other multidomain allosteric proteins.

biophysics↗

bioRxiv2023

PMS1 as a target for splice modulation to prevent somatic CAG repeat expansion in Huntington's disease

Huntingtons disease (HD) is a dominantly inherited neurodegenerative disorder whose motor, cognitive, and behavioral manifestations are caused by an expanded, somatically unstable CAG repeat in the first exon of HTT that lengthens a polyglutamine tract in huntingtin. Genome-wide association studies (GWAS) have revealed DNA repair genes that influence the age-at-onset of HD and implicate somatic CAG repeat expansion as the primary driver of disease timing. To prevent the consequent neuronal damage, small molecule splice modulators (e.g., branaplam) that target HTT to reduce the levels of huntingtin are being investigated as potential HD therapeutics. We found that the effectiveness of the splice modulators can be influenced by genetic variants, both at HTT and other genes where they promote pseudoexon inclusion. Surprisingly, in a novel hTERT-immortalized retinal pigment epithelial cell (RPE1) model for assessing CAG repeat instability, these drugs also reduced the rate of HTT CAG expansion. We determined that the splice modulators also affect the expression of the mismatch repair gene PMS1, a known modifier of HD age-at-onset. Genome editing at specific HTT and PMS1 sequences using CRISPR-Cas9 nuclease confirmed that branaplam suppresses CAG expansion by promoting the inclusion of a pseudoexon in PMS1, making splice modulation of PMS1 a potential strategy for delaying HD onset. Comparison with another splice modulator, risdiplam, suggests that other genes affected by these splice modulators also influence CAG instability and might provide additional therapeutic targets.

genetics↗

bioRxiv2022

A naturally occurring lytic bioprocess improves sustainability of phycocyanin production from cyanobacteria

Cyanobacteria form dense multicellular communities that experience transient conditions in terms of access to light and oxygen. These systems are productive but also undergo substantial biomass turnover, supplementing heightened heterotrophic respiration and oxygen drawdown. Here we use metagenomics and metaproteomics to survey the cellular response of a mat-forming cyanobacterium undergoing mass cell lysis after exposure to dark and anoxic conditions. A lack of evidence for visral, bacterial, or eukaryotic antagonism contradicts commonly held beliefs on the causative agent for cyanobacterial death during dense growth. Instead, proteogenomics data indicated that lysis resulted from a genetically programmed response triggered by a failure to maintain osmotic pressure in the wake of severe energy limitation. Cyanobacterial DNA was rapidly degraded, yet cyanobacterial proteins remained abundant. A subset of proteins, including enzymes involved in amino acid metabolism, peptidases, toxin-antitoxin systems, and a potentially self-targeting CRISPR-Cas system, were upregulated upon lysis, indicating involvement in the programmed cell death response. We propose this natural form of programmed cell death could provide new pathways for controlling harmful algal blooms and for sustainable bioproduct production.

microbiology↗

bioRxiv2022

Tissue-specific and repeat length-dependent somatic instability of the X-linked dystonia parkinsonism-associated CCCTCT repeat

X-linked dystonia-parkinsonism (XDP) is a progressive adult-onset neurodegenerative disorder caused by insertion of a SINE-VNTR-Alu (SVA) retrotransposon in the TAF1 gene. The SVA retrotransposon contains a CCCTCT hexameric repeat tract of variable length, whose length is inversely correlated with age at onset. This places XDP in a broader class of repeat expansion diseases, characterized by the instability of their causative repeat mutations. Here, we observe similar inverse correlations between CCCTCT repeat length with age at onset and age at death and no obvious correlation with disease duration. To gain insight into repeat instability in XDP we performed comprehensive quantitative analyses of somatic instability of the XDP CCCTCT repeat in blood and in seventeen brain regions from affected males. Our findings reveal repeat length-dependent and expansion-based instability of the XDP CCCTCT repeat, with greater levels of expansion in brain than in blood. The brain exhibits regional-specific patterns of instability that are broadly similar across individuals, with cerebellum exhibiting low instability and cortical regions exhibiting relatively high instability. The spectrum of somatic instability in the brain includes a high proportion of moderate repeat length changes of up to 5 repeats, as well as expansions of [~]20->100 repeats and contractions of [~]20-40 repeats at lower frequencies. Comparison with HTT CAG repeat instability in postmortem Huntingtons disease brains reveals similar brain region-specific profiles, indicating common trans-acting factors that contribute to the instability of both repeats. Analyses in XDP brains of expansion of a different SVA-associated CCCTCT located in the LIPG gene, and not known to be disease-associated, reveals repeat length-dependent expansion at overall lower levels relative to the XDP CCCTCT repeat, suggesting that expansion propensity may be modified by local chromatin structure. Together, the data support a role for repeat length-dependent somatic expansion in the process(es) driving the onset of XDP and prompt further investigation into repeat dynamics and the relationship to disease.

genetics↗

bioRxiv2019

Genetic overlap between psychiatric disorders and neuropsychiatric symptoms in HD

Huntingtons disease (HD) is an inherited neurodegenerative disorder caused by an expanded CAG repeat in the HTT gene. It is diagnosed following a standardized exam of motor control and often presents with cognitive decline and psychiatric symptoms. Recent studies have detected genetic loci modifying the age at onset of motor symptoms in HD, but genetic factors influencing cognitive and psychiatric presentations are unknown. We tested the hypothesis that psychiatric and cognitive symptoms in HD are influenced by the same common genetic variation as in the general population by constructing polygenic risk scores from large genome-wide association studies of psychiatric and neurodegenerative disorders, and of intelligence, and testing for correlation with the presence of psychiatric and cognitive symptoms in a large sample (n=5160) of HD patients. Polygenic risk score for major depression was associated specifically with increased risk of depression in HD, as was schizophrenia risk score with psychosis and irritability. Cognitive impairment and apathy were associated with reduced polygenic risk score for intelligence. In general, polygenic risk scores for psychiatric disorders, particularly depression and schizophrenia, are associated with increased risk of the corresponding psychiatric symptoms in HD, suggesting a common genetic liability. However, the genetic liability to cognitive impairment and apathy appears to be distinct from other psychiatric symptoms in HD. No associations were observed between HD symptoms and risk scores for other neurodegenerative disorders. These data provide a rationale for treatments effective in depression and schizophrenia to be used to treat depression and psychotic symptoms in HD.

genetics↗

bioRxiv2019

Huntington's disease onset is determined by length of uninterrupted CAG, not encoded polyglutamine, and is modified by DNA maintenance mechanisms

The effects of variable, glutamine-encoding, CAA interruptions indicate that a property of the uninterrupted HTT CAG repeat sequence, distinct from huntingtins polyglutamine segment, dictates the rate at which HD develops. The timing of onset shows no significant association with HTT cis-eQTLs but is influenced, sometimes in a sex-specific manner, by polymorphic variation at multiple DNA maintenance genes, suggesting that the special onset-determining property of the uninterrupted CAG repeat is a propensity for length instability that leads to its somatic expansion. Additional naturally-occurring genetic modifier loci, defined by GWAS, may influence HD pathogenesis through other mechanisms. These findings have profound implications for the pathogenesis of HD and other repeat diseases and question a fundamental premise of the "polyglutamine disorders".

genetics↗