Biology subjects

Kanai, M.

Publications and source records attributed to Kanai, M..

At least 19 recordsLinked to original sources

bioRxiv2026

sc-pcQTL: hurdle-based co-expression modeling for multi-gene QTL mapping in single-cell RNA-seq data

Motivation: Single-cell expression quantitative trait locus (eQTL) studies can resolve cell-type-specific genetic effects, but conventional gene-by-gene analyses do not directly capture coordinated genetic regulation of neighboring genes. Principal-component QTL (pcQTL) mapping can summarize such multi-gene effects, but existing approaches were developed for bulk expression and are not designed for sparse single-cell counts. Results: We developed sc-pcQTL, a framework that applies two-component hurdle modeling and sliding-window clustering to identify local co-expression clusters, summarizes each cluster using principal components, and maps cis-pcQTLs. In simulations, the individual hurdle components controlled type I error, while the component-union screening rule was substantially more powerful than donor-level pseudobulk correlation tests. Applied to 1.24 million peripheral blood mononuclear cells from 982 OneK1K donors across 10 cell types, sc-pcQTL identified 2,485 local co-expression clusters and conducted QTL mapping for 4,353 cluster-PC phenotypes at single-cell resolution, of which 2,040 had at least one significant cis-pcQTL association. Fine-mapping and colocalization with genome-wide association study loci across 1,163 phenotypes in the FinnGen study identified 394 colocalized QTL-GWAS signal groups. Each group comprised fine-mapped QTL and GWAS signals connected through one or more colocalization links within the same cell type and local gene cluster. Of these groups, 46 were pcQTL-specific and contained no colocalized single-gene eQTL from a constituent gene. Locus-level analyses further revealed cell-type-specific multi-gene regulatory effects. Thus, sc-pcQTL complements conventional single-gene eQTL analysis by identifying trait-relevant regulatory signals shared across neighboring genes.

bioinformatics↗

bioRxiv2025

The complete mitochondrial genomes of Litophyton sp. and Stereonephthya sp., members of family Nephtheidae (Cnidaria: Anthozoa: Octocorallia).

In this study, we sequenced the complete mitochondrial genomes (mitogenome) of Litophyton sp. and Stereonephthya sp. (family Nephtheidae), collected in Okinawa, Japan. The complete mitogenomes of Litophyton sp. and Stereonephthya sp. were 19,130 bp and 18,912 bp in length, respectively. They contained 17 genes (14 protein-coding genes, two rRNA genes, and one tRNA gene) and exhibited gene order pattern "A", which is the most common arrangement among octocorals. Molecular phylogenetic analysis indicated that Litophyton sp. And Stereonephthya sp. are clustered with other species belonging to family Nephtheidae, consistent with the previous research using partial mitochondrial genes. Our study provides additional mitochondrial genomic resources that contributes to comparative studies of octocoral mitogenomes.

genomics↗

bioRxiv2025

The complete mitochondrial genome of Hanabira yukibana Lau, Stokvis, Imahara & Reimer, 2019 (Cnidaria: Anthozoa: Octocorallia)

In this study, we sequenced the complete mitochondrial genome (mitogenome) of Hanabira yukibana (family Clavulariidae), collected in Okinawa, Japan. The complete mitochondrial genome of this specimen was 18,690 bp in length and contained 17 genes (14 protein-coding genes, two rRNA genes, and one tRNA gene). It exhibited gene order pattern A, which is the most common arrangement among octocorals. Molecular phylogenetic analysis indicated that H. yukibana is clustered with the genus Clavularia, including Clavularia inflata (the family Clavulariidae), consistent with the previous research using partial mitochondrial genes. Our study provides an additional mitochondrial genomic resource that contributes to comparative studies of octocoral mitogenomes.

genomics↗

bioRxiv2025

The mitochondrial genome of Palythoa tuberculosa (Esper, 1805) contains a novel ORF with unknown function.

Recent advances in sequencing technology have enabled rapid and cost-effective recovery of mitochondrial genomes (mt-genomes), which used in phylogenetic and evolutionary studies. Among cnidarians, the order Zoantharia possesses the largest mt-genomes on average (20,976 bp). Here, we present the complete mt-genome of Palythoa tuberculosa (Hexacorallia: Zoantharia: Sphenopidae), which was 23,044 bp in length--approximately 2,000 bp larger than the zoantharian average. Synteny analysis revealed that this increase resulted from a novel open reading frame (ORF) located between trnW and cox2. This ORF was also present in the closely related species P. caribaeorum, indicating its origin in their common ancestor. Importantly, this represents the first case of a novel ORF insertion in zoantharians, clearly distinct from previously reported ORF extensions. While mt-genome expansions are occasionally observed in cnidarians, both the insertion sites and inserted elements vary among lineages, highlighting the lineage-specific mechanisms driving mitochondrial genome evolution in this group.

genomics↗

bioRxiv2025

Sensitive Transcriptomics and Genotyping reveals function of genetic variants in immunity

Advancements in genomics have revolutionized human genetics by defining the genetic architecture of disease. However, identifying causal variants and their mechanisms of action remains a challenge in translating genetics into therapeutic interventions. Here, we developed Sensitive Transcriptomics And Genotyping by sequencing (STAG-seq), a high-throughput platform designed to define mechanistic genotype-phenotype relationships through simultaneous single-cell measurements of genomic DNA and RNA transcripts. Combined with base-editing, STAG-seq enables functionalization of variants in relevant cellular contexts. We demonstrate the applicability of this approach in several settings. First, we screened genetic perturbations to identify monoallelic and biallelic variant effects in primary human macrophages treated with innate immune stimuli. Next, we phenotyped clinically relevant missense variants associated with immunodeficiency and autoimmunity. Finally, we defined a noncoding variant in a pleiotropic autoimmunity locus that governs TNRC18 expression in primary T cells. STAG-seq thus enables variant phenotyping at scale to advance functional genomics and disease biology.

genomics↗

bioRxiv2024

Identification of the drug/metabolite transporter 1 as a marker of quinine resistance in a NF54xCam3.II P. falciparum genetic cross.

The genetic basis of Plasmodium falciparum resistance to quinine (QN), a drug used to treat severe malaria, has long been enigmatic. To gain further insight, we used FRG-NOD human liver-chimeric mice to conduct a P. falciparum genetic cross between QN-resistant (Cam3.II) and QN-sensitive (NF54) parasites, which also differ in their susceptibility to chloroquine (CQ). By applying different selective conditions to progeny pools prior to cloning, we recovered 120 unique recombinant progeny. Drug profiling and quantitative trait loci analyses of the progeny revealed predominant peaks on chromosomes 7 and 12 associated with CQ and QN resistance, that is consistent with a multifactorial mechanism of resistance for these compounds. CQ and monodesethyl-CQ (md-CQ) resistance mapped to a chromosome 7 region harboring pfcrt as expected. However, for QN, resistance mapped to a dominant chromosome 7 peak centered 295 kb downstream of pfcrt, with pfcrt showing a smaller peak. We identified the drug/metabolite transporter 1 (DMT1) as the top chromosome 7 candidate due to its structural similarity to PfCRT and proximity to the peak. Deleting DMT1 in QN-resistant Cam3.II parasites significantly sensitized the parasite to QN but not to the other drugs tested, suggesting that DMT1 mediates QN response specifically. We localized DMT1 to structures associated with vesicular trafficking, as well as the parasitophorous vacuolar membrane, lipid bodies, and the digestive vacuole. We also observed that mutant DMT1 transports more QN than the wild-type isoform in vitro. Gene editing confirmed an additional role for mutant PfCRT in mediating QN resistance. In addition, we identified an ATP-dependent zinc metalloprotease (FtsH1) as one of the top candidates in the chromosome 12 locus and confirmed its role as a potential mediator of QN resistance and a modulator of md-CQ resistance using CRISPR/Cas9 SNP-edited lines. Interestingly, this chromosome 12 region mapped to resistance to both CQ and QN and was preferentially co-inherited with pfcrt. Our study demonstrates that DMT1 is a novel marker of QN resistance and that a new chromosome 12 locus associates with CQ and QN response, with ftsh1 as a potential candidate, suggesting these genes in addition to pfcrt should be genotyped in surveillance and clinical settings.

genetics↗

bioRxiv2024

Variant scoring performance across selection regimes depends on variant-to-gene and gene-to-disease components

Variant scoring methods (VSMs) aid in the interpretation of coding mutations and their potential impact on health, but their evaluation in the context of human genetics applications remains inconsistent. Here, we describe GeneticsGym, a systematic approach to evaluating the real-world impact of VSMs on human genetic analysis. We show that the relative performance of VSMs varies across regimes of natural selection, and that both variant-to-gene and gene-to-disease components contribute.

bioinformatics↗

bioRxiv2024

Widespread naturally variable human exons aid genetic interpretation

Most mammalian genes undergo alternative splicing. The splicing of some exons has been acquired or lost in specific mammalian lineages, but differences in splicing within the human population are poorly characterized. Using GTEx tissue transcriptomes from 838 individuals, we identified 56,415 exons which are included in mRNAs in some individuals but entirely excluded from others, which we term "naturally variable exons" (NVEs). NVEs impact three quarters of protein-coding genes, occur at all population frequencies, and are often absent from reference annotations. NVEs are more abundant in genes depleted of genetic loss-of-function mutations and aid in the interpretation of causal genetic variants. Genetic variants modulate the splicing of many NVEs, and 5UTR and coding-region NVEs are often associated with increased and decreased gene expression, respectively. Together, our findings characterize abundant splicing variation in the human population, with implications for a range of human genetic analyses.

genetics↗

bioRxiv2024

Functional dissection of complex and molecular trait variants at single nucleotide resolution

Identifying the causal variants and mechanisms that drive complex traits and diseases remains a core problem in human genetics. The majority of these variants have individually weak effects and lie in non-coding gene-regulatory elements where we lack a complete understanding of how single nucleotide alterations modulate transcriptional processes to affect human phenotypes. To address this, we measured the activity of 221,412 trait-associated variants that had been statistically fine-mapped using a Massively Parallel Reporter Assay (MPRA) in 5 diverse cell-types. We show that MPRA is able to discriminate between likely causal variants and controls, identifying 12,025 regulatory variants with high precision. Although the effects of these variants largely agree with orthogonal measures of function, only 69% can plausibly be explained by the disruption of a known transcription factor (TF) binding motif. We dissect the mechanisms of 136 variants using saturation mutagenesis and assign impacted TFs for 91% of variants without a clear canonical mechanism. Finally, we provide evidence that epistasis is prevalent for variants in close proximity and identify multiple functional variants on the same haplotype at a small, but important, subset of trait-associated loci. Overall, our study provides a systematic functional characterization of likely causal common variants underlying complex and molecular human traits, enabling new insights into the regulatory grammar underlying disease risk.

genetics↗

bioRxiv2023

Dual-specificity phosphatases 13 and 27 as key switches in muscle stem cell transition from proliferation to differentiation

Muscle regeneration depends on muscle stem cell (MuSC) activity. Myogenic regulatory factors, including myoblast determination protein 1 (MyoD), regulate the fate transition of MuSCs. However, the direct target of MYOD in the process is not completely clear. Using previously established MyoD knock-in (MyoD-KI) mice, we revealed that MyoD targets dual-specificity phosphatase (Dusp) 13 and Dusp27. In Dusp13:Dusp27 double knock-out (DKO) mice, the ability for muscle regeneration after injury was reduced. Moreover, single-cell RNA sequencing of MyoD-high expressing MuSCs from MyoD-KI mice revealed that Dusp13 and Dusp27 are expressed only in specific populations within MyoD-high MuSCs, which also express Myogenin. Overexpressing Dusp13 in MuSCs causes premature muscle differentiation. Thus, we propose a model where DUSP13 and DUSP27 contribute to the fate transition of MuSCs from proliferation to differentiation during myogenesis. Significance StatementMYOD protein is not expressed in quiescent muscle stem cells but accumulates rapidly following muscle injury, leading to the proliferation of myogenic progenitors for differentiation. However, the direct targets of MYOD, aside from myogenin, which play roles in myogenic differentiation remain incompletely understood. Using previously established MyoD knock-in mice and single-cell RNA sequencing, we discovered that Dusp13 and Dusp27 are potential target genes of MYOD that promote myogenesis during muscle regeneration in adult mice.

cell biology↗

bioRxiv2023

Mapping the genomic landscape of multidrug resistance in Plasmodium falciparum and its impact on parasite fitness

Drug-resistant Plasmodium falciparum parasites have swept across Southeast Asia and now threaten Africa. By implementing a P. falciparum genetic cross using humanized mice, we report the identification of key determinants of resistance to artemisinin (ART) and piperaquine (PPQ) in the dominant Asian KEL1/PLA1 lineage. We mapped k13 as the central mediator of ART resistance and identified secondary markers. Applying bulk segregant analysis, quantitative trait loci mapping and gene editing, our data reveal an epistatic interaction between mutant PfCRT and multicopy plasmepsins 2/3 in mediating high-grade PPQ resistance. Susceptibility and parasite fitness assays implicate PPQ as a driver of selection for KEL1/PLA1 parasites. Mutant PfCRT enhanced susceptibility to lumefantrine, the first-line partner drug in Africa, highlighting a potential benefit of opposing selective pressures with this drug and PPQ. We also identified that the ABCI3 transporter can operate in concert with PfCRT and plasmepsins 2/3 in mediating multigenic resistance to antimalarial agents.

genomics↗

bioRxiv2023

Systematic visualisation of molecular QTLs reveals variant mechanisms at GWAS loci

Splicing quantitative trait loci (QTLs) have been implicated as a common mechanism underlying complex trait associations. However, utilising splicing QTLs in target discovery and prioritisation has been challenging due to extensive data normalisation which often renders the direction of the genetic effect as well as its magnitude difficult to interpret. This is further complicated by the fact that strong expression QTLs often manifest as weak splicing QTLs and vice versa, making it difficult to uniquely identify the underlying molecular mechanism at each locus. We find that these ambiguities can be mitigated by visualising the association between the genotype and average RNA sequencing read coverage in the region. Here, we generate these QTL coverage plots for 1.7 million molecular QTL associations in the eQTL Catalogue identified with five quantification methods. We illustrate the utility of these QTL coverage plots by performing colocalisation between vitamin D levels in the UK Biobank and all molecular QTLs in the eQTL Catalogue. We find that while visually confirmed splicing QTLs explain just 6/53 of the colocalising signals, they are significantly less pleiotropic than eQTLs and identify a prioritised causal gene in 4/6 cases. All our association summary statistics and QTL coverage plots are freely available at https://www.ebi.ac.uk/eqtl/.

genomics↗

bioRxiv2022

Polygenic prediction across populations is influenced by ancestry, genetic architecture, and methodology

Polygenic risk scores (PRS) developed from multi-ancestry genome-wide association studies (GWAS), PRSmulti, hold promise for improving PRS accuracy and generalizability across populations. To establish best practices for leveraging the increasing diversity of genomic studies, we investigated how various factors affect the performance of PRSmulti compared to PRS constructed from single-ancestry GWAS (PRSsingle). Through extensive simulations and empirical analyses, we showed that PRSmulti overall outperformed PRSsingle in understudied populations, except when the understudied population represented a small proportion of the multi-ancestry GWAS. Notably, for traits with large-effect ancestry-enriched variants, such as mean corpuscular volume, using substantially fewer samples from Biobank Japan achieved comparable accuracies to a much larger European cohort. Furthermore, integrating PRS based on local ancestry-informed GWAS and large-scale European-based PRS improved predictive performance in understudied African populations, especially for less polygenic traits with large ancestry-enriched variants. Our work highlights the importance of diversifying genomic studies to achieve equitable PRS performance across ancestral populations and provides guidance for developing PRS from multiple studies.

genomics↗

bioRxiv2022

Actin filaments accumulate in the nucleus during interphase and remain in the vicinity of condensing chromosomes during prophase to metaphase in zebrafish early embryos

In the cytoplasm, filamentous actin (F-actin) plays a critical role in cell regulation, including cell migration, stress fiber formation, and cytokinesis. Recent studies have shown that actin filaments that form in the nucleus are associated with diverse functions. Here, using live imaging of an F-actin-specific probe, superfolder GFP-tagged utrophin (UtrCH-sfGFP), we demonstrated the dynamics of nuclear actin in zebrafish (Danio rerio) embryos. In early zebrafish embryos up to around the high stage, UtrCH-sfGFP increasingly accumulated in nuclei during the interphase and reached a peak during the prophase. After nuclear envelope breakdown (NEBD), patches of UtrCH-sfGFP remained in the vicinity of condensing chromosomes during the prometaphase to metaphase. When zygotic transcription was inhibited by injecting -amanitin, the nuclear accumulation of UtrCH-sfGFP was still observed at the sphere and dome stages, suggesting that zygotic transcription may induce a decrease in nuclear F-actin. The accumulation of F-actin in nuclei may contribute to proper mitotic progression of large cells with rapid cell cycles in zebrafish early embryos, by assisting in NEBD, chromosome congression, and/or spindle assembly. Summary statementFilamentous actin accumulates in the nucleus of zebrafish early embryos and forms patches associating with condensing chromosomes during prophase.

cell biology↗

bioRxiv2022

Improving fine-mapping by modeling infinitesimal effects

Fine-mapping aims to identify causal variants for phenotypes. Bayesian fine-mapping algorithms (e.g.: SuSiE, FINEMAP, ABF, and COJO-ABF) are widely used, but assessing posterior probability calibration remains challenging in real data, where model misspecification likely exists, and true causal variants are unknown. We introduce Replication Failure Rate (RFR), a metric to assess fine-mapping consistency by down-sampling. SuSiE, FINEMAP and COJO-ABF show high RFR, indicating potential under-conservative mis-calibration. Simulations reveal that non-sparse genetic architecture can lead to miscalibration, while imputation noise, non-uniform distribution of causal variants, and QC filters have minimal impact. We present SuSiE-inf and FINEMAP-inf, novel fine-mapping methods modeling infinitesimal effects alongside fewer larger causal effects. Our methods exhibit improved calibration, RFR and functional enrichment, competitive recall and computational efficiency. Notably, using our methods posterior effect sizes substantially increases PRS accuracy over SuSiE and FINEMAP. Our work improves causal variants identification for complex traits, a fundamental goal of human genetics.

bioinformatics↗

bioRxiv2022

Modulation of the functional interfaces between retroviral intasomes and the human nucleosome

Retroviral integration into cell chromatin requires the formation of integrase-viral DNA complexes, called intasomes, and their interaction with the target DNA wrapped around nucleosomes. To further study this mechanism we developed an alphaLISA approach using the prototype foamy virus (PFV) intasome and human nucleosome. This system allowed us to monitor the association between both partners and investigate the protein/protein and protein/DNA interactions engaged in the association with chromatin. Using this approach, we next screened the chemical OncoSET library and selected small molecules that could modulate the intasome/nucleosome complex. Molecules were selected as acting either on the DNA topology within the nucleosome or on the integrase/histone tail interactions. Within these compounds, doxorubicin and histone binders calixarenes were characterized using biochemical, structural and cellular approaches. These drugs were shown to inhibit PFV and HIV-1 integration in vitro as well as HIV-1 infection in primary PBMCs cells. Our work provides new information about intasome-nucleosome interaction determinants and paves the way for further unedited antiviral strategies that target the final step of intasome/chromatin anchoring.

microbiology↗

bioRxiv2022

Multi-ancestry GWAS of major depression aids locus discovery, fine-mapping, gene prioritisation, and causal inference

Most genome-wide association studies (GWAS) of major depression (MD) have been conducted in samples of European ancestry. Here we report a multi-ancestry GWAS of MD, adding data from 21 studies with 88,316 MD cases and 902,757 controls to previously reported data from individuals of European ancestry. This includes samples of African (36% of effective sample size), East Asian (26%) and South Asian (6%) ancestry and Hispanic/Latinx participants (32%). The multi-ancestry GWAS identified 190 significantly associated loci, 53 of them novel. For previously reported loci from GWAS in European ancestry the power-adjusted transferability ratio was 0.6 in the Hispanic/Latinx group and 0.3 in each of the other groups. Fine-mapping benefited from additional sample diversity: the number of credible sets with [≤]5 variants increased from 3 to 12. A transcriptome-wide association study identified 354 significantly associated genes, 205 of them novel. Mendelian Randomisation showed a bidirectional relationship with BMI exclusively in samples of European ancestry. This first multi-ancestry GWAS of MD demonstrates the importance of large diverse samples for the identification of target genes and putative mechanisms.

genetics↗

bioRxiv2021

A multi-layer functional genomic analysis to understand noncoding genetic variation in lipids

A major challenge of genome-wide association studies (GWAS) is to translate phenotypic associations into biological insights. Here, we integrate a large GWAS on blood lipids involving 1.6 million individuals from five ancestries with a wide array of functional genomic datasets to discover regulatory mechanisms underlying lipid associations. We first prioritize lipid-associated genes with expression quantitative trait locus (eQTL) colocalizations, and then add chromatin interaction data to narrow the search for functional genes. Polygenic enrichment analysis across 697 annotations from a host of tissues and cell types confirms the central role of the liver in lipid levels, and highlights the selective enrichment of adipose-specific chromatin marks in high-density lipoprotein cholesterol and triglycerides. Overlapping transcription factor (TF) binding sites with lipid-associated loci identifies TFs relevant in lipid biology. In addition, we present an integrative framework to prioritize causal variants at GWAS loci, producing a comprehensive list of candidate causal genes and variants with multiple layers of functional evidence. Two prioritized genes, CREBRF and RRBP1, show convergent evidence across functional datasets supporting their roles in lipid biology.

genomics↗