Biology subjects

Muratani, M.

Publications and source records attributed to Muratani, M..

5 recordsLinked to original sources

bioRxiv2025

Comprehensive and accelerated mapping of driver mutations through single-nucleotide random mutagenesis of target genes

Identification of novel driver mutations is crucial for personalized medicine and drug discovery. However, genome-wide studies based solely on patient-derived data cannot confirm the oncogenic potential of detected mutations. Here, we present a genome editing-based method for comprehensive random mutagenesis of target genes. This approach generates cultured cells harboring random single-nucleotide variants (SNVs) within 10 days of DNA amplicon preparation. To validate its utility, random mutations were introduced into endogenous exons of lung cancer-related oncogenes in HEK293 cells. Spheroid formation assays and xenograft models demonstrated enhanced tumorigenicity, indicating the acquisition of oncogenic potential. Amplicon-sequencing of spheroid transcripts revealed significant enrichment of variants annotated as pathogenic in clinical databases and computational predictions. Furthermore, osimertinib treatment of lung cancer cell lines with random SNVs on EGFR identified 58 novel resistance mutations. These results highlight the effectiveness of our mutagenesis platform, offering a powerful tool to advance cancer research and precision medicine.

bioengineering↗

bioRxiv2024

Large MAF Transcription Factors Reawaken Evolutionarily Dormant Fast-Glycolytic Type IIb Myofibers in Human Skeletal Muscle

Small mammals rely on type IIb myofibers, expressing the fastest myosin IIb (encoded by MYH4), for rapid muscle contraction. In contrast, larger mammals, including humans, show reduced or absent MYH4 expression and type IIb myofibers, favoring slower-contracting myofibers. The evolutionary mechanisms underlying this shift remain unclear. Here, we identify large MAF transcription factors (MAFA, MAFB, MAF) as key regulators of MYH4 expression in large mammals, including human and bovine. Overexpression of large MAFs induces MYH4 expression and enhances glycolytic capacity in human myotubes, supported by RNA-seq and metabolic flux analyses. RNA-seq of human muscle biopsies reveals a positive correlation between MAFA, MAF, and MYH4 expression, with these genes elevated in power-trained individuals. These findings reveal a conserved mechanism across mammals, showing that large MAFs can induce type IIb myofibers even in humans, with potential applications for enhancing athletic performance and addressing age-related muscle weakness associated with the loss of fast-twitch myofibers.

cell biology↗

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↗

bioRxiv2020

Transcriptome analysis of sevoflurane exposure effects at the different brain regions

BackgroundsSevoflurane is a most frequently used volatile anaesthetics, but its molecular mechanisms of action remain unclear. We hypothesized that specific genes play regulatory roles in whole brain exposed to sevoflurane. Thus, we aimed to evaluate the effects of sevoflurane inhalation and identify potential regulatory genes by RNA-seq analysis. MethodsEight-week old mice were exposed to sevoflurane. RNA from four medial prefrontal cortex, striatum, hypothalamus, and hippocampus were analysed using RNA-seq. Differently expressed genes were extracted. Their gene ontology terms and the transcriptome array data of the cerebral cortex of sleeping mice were analysed using Metascape, and the gene expression patterns were compared. Finally, the activities of transcription factors were evaluated using a weighted parametric gene set analysis (wPGSA). JASPAR was used to confirm the existence of binding motifs in the upstream sequences of the differently expressed genes. ResultsThe gene ontology term enrichment analysis result suggests that sevoflurane inhalation upregulated angiogenesis and downregulated neural differentiation in the whole brain. The comparison with the brains of sleeping mice showed that the gene expression changes were specific to anaesthetized mice. Sevoflurane induced Klf4 upregulation in the whole brain. The transcriptional analysis result suggests that KLF4 is a potential transcriptional regulator of angiogenesis and neural development. ConclusionsKlf4 was upregulated by sevoflurane inhalation in whole brain. KLF4 might promote angiogenesis and cause the appearance of undifferentiated neural cells by transcriptional regulation. The roles of KLF4 might be key to elucidating the mechanisms of sevoflurane induced functional modification in the brain.

genetics↗

bioRxiv2019

Cables2 Is a novel Smad2-regulatory factor essential for early embryonic development in mice

CDK5 and Abl enzyme substrate 2 (Cables2), a member of the Cables family that has a C-terminal cyclin box-like domain, is widely expressed in adult mouse tissues. However, the physiological role of Cables2 in vivo is unknown. We show here that Cables2-deficiency causes post-gastrulation embryonic lethality in mice. The mutant embryos progress to gastrulation, but then arrest, and fail to grow. Analysis of gene expression patterns reveals that formation of the anterior visceral endoderm and the primitive streak is impaired in Cables2-deficient embryos. Tetraploid complementation analyses support the critical requirement of Cables2 in both the epiblast and visceral endoderm for progression of embryogenesis. In addition, we show that Cables2 physically interacts with a key mediator of the canonical Nodal pathway, Smad2, and augments its transcriptional activity. These findings provide novel insights into the essential role of Cables2 in the early embryonic development in mice.

developmental biology↗