Biology subjects

Shimizu, C.

Publications and source records attributed to Shimizu, C..

9 recordsLinked to original sources

bioRxiv2026

SIK3 links synaptic strength to sleep pressure through RhoA signaling

Sleep is regulated by homeostatic sleep pressure, which accumulates during wakefulness, triggering sleep, and dissipates during sleep, leading to wakefulness. Although recent evidence suggests that synaptic potentiation encodes sleep pressure, the intracellular mechanisms linking synaptic strength to sleep pressure remain unclear. Here we demonstrate that the sleep-promoting kinase SIK3 links synaptic strength and sleep pressure through RhoA signalling. We found that SIK3 is required for dendritic spine enlargement induced by chemical long-term potentiation (cLTP) in cultured cortical pyramidal neurons and positively regulates spine size in the prefrontal cortex in vivo. cLTP reduced RhoA activity in an SIK3-dependent manner during spine enlargement, whereas activation of RhoA reduced EEG delta density in vivo during non-rapid eye movement sleep (NREMS), an indicator of sleep pressure. SIK3 substrate screening identified several RhoA regulators including GEF-H1, and further biochemical analyses found that phosphorylation by SIK3 activates GEF-H1. However, contrary to our expectation, RhoA facilitation through pharmacological activation of GEF-H1 enlarged spine size in cultured neurons, and expression of constitutively active GEF-H1 increased NREMS delta density in a RhoA-dependent fashion. Together, these results indicate that SIK3 enlarges spine size and enhances NREMS delta density through two RhoA-dependent pathways, driving mutually opposing intermediate phenotypes. Our findings identify two clashing SIK3-RhoA signalling pathways that couple synaptic regulation to sleep homeostasis.

neuroscience↗

bioRxiv2026

Reactivating a schizophrenia risk gene-enriched prefrontal ensemble suppresses decision noise

Schizophrenia genetics and single-cell transcriptomics implicate prefrontal excitatory neurons and synaptic programs, but how these vulnerabilities alter circuit dynamics, behavior, and pharmacological targetability remains unclear. We identify a clozapine-responsive prefrontal ensemble enriched for schizophrenia risk genes that stabilizes value-guided choice. NMDA receptor hypofunction increased value-independent decision noise and weakened the ensembles pre-choice transient, while clozapine rescued both. Chemogenetic inhibition of the ensemble increased decision noise, demonstrating causality. By integrating receptor affinities with mouse and human single-cell transcriptomes, we designed a rational multi-receptor antagonist cocktail that reactivated the ensemble and rescued decision noise without increasing NREM sleep time or NREM delta power, in contrast to clozapine. These findings link schizophrenia genetics to cortical ensemble dynamics and establish ensemble-targeted pharmacology as a strategy for ameliorating pathological computations. One-Sentence SummaryTargeting a schizophrenia risk gene-enriched prefrontal ensemble suppresses decision noise without clozapine-like sedation.

neuroscience↗

bioRxiv2025

Minimal Correlation but Complementary Diagnostic Utility for Plasma Cell-free RNA and Proteins

Proteins and RNA circulate in plasma and can offer insights into human physiology. Yet, despite their clinical importance, direct comparisons between these analytes remain unexplored. Here, we measured and compared plasma cell-free RNA (cfRNA) and protein levels for 263 children diagnosed with inflammatory diseases by RNA-sequencing (n=155) and SomaScan proteomics (n=171). Remarkably, cfRNA and protein levels were largely uncorrelated across samples (feature-by-sample r=0.052; median feature-level r=0.009). Nonetheless, machine learning models based on either modality distinguished Kawasaki Disease (KD) from Multisystem Inflammatory Syndrome in Children (MIS-C) with similar high accuracy (median AUC > 0.93). Analysis of KD subtypes revealed distinct cfRNA and protein signatures, with one group showing molecular similarity to MIS-C. These findings underscore the complementary nature of cfRNA and protein profiling and highlight the utility of integrating multiple blood analytes to improve disease classification and deepen our understanding of complex inflammatory conditions.

genomics↗

bioRxiv2023

Cortical parvalbumin neurons are responsible for homeostatic sleep rebound through CaMKII activation

The homeostatic regulation of sleep is characterized by rebound sleep after prolonged wakefulness, but the molecular and cellular mechanisms underlying this regulation are still unknown. We show here that CaMKII-dependent activity control of parvalbumin (PV)-expressing cortical neurons is involved in sleep homeostasis regulation. Prolonged wakefulness enhances cortical PV-neuron activity. Chemogenetic suppression or activation of cortical PV neurons inhibits or induces rebound sleep, implying that rebound sleep is dependent on increased activity of cortical PV neurons. Furthermore, we discovered that CaMKII kinase activity boosts the activity of cortical PV neurons, and that kinase activity is important for homeostatic sleep rebound. We propose that CaMKII-dependent PV-neuron activity represents negative feedback inhibition of cortical neural excitability, which serves as the distributive cortical circuits for sleep homeostatic regulation.

neuroscience↗

bioRxiv2023

Bioorthogonal chemical labelling of endogenous neurotransmitter receptors in living mouse brains

Neurotransmitter receptors are essential components of synapses for communication between neurons in the brain. Because the spatiotemporal expression profiles and dynamics of neurotransmitter receptors involved in many functions are delicately governed in the brain, in vivo research tools with high spatiotemporal resolution for receptors in intact brains are highly desirable. Covalent chemical labelling of proteins without genetic manipulation is now a powerful method for analyzing receptors in vitro. However, selective target receptor labelling in the brain has not yet been achieved. This study shows that ligand-directed alkoxyacylimidazole (LDAI) chemistry can be used to selectively tether synthetic probes to target endogenous receptors in living mouse brains. The reactive LDAI reagents with negative charges were found to diffuse well over the whole brain and could selectively label target endogenous receptors, including AMPAR, NMDAR, mGlu1, and GABAAR. This simple and robust labelling protocol was then used for various applications: three-dimensional spatial mapping of endogenous receptors in the brains of healthy and disease-model mice; multi-colour receptor imaging; and pulse-chase analysis of the receptor dynamics in postnatal mouse brains. Here, results demonstrated that bioorthogonal receptor modification in living animal brains may provide innovative molecular tools that contribute to the in-depth understanding of complicated brain functions.

biochemistry↗

bioRxiv2022

Autoantibody discovery across monogenic, acquired, and COVID19-associated autoimmunity with scalable PhIP-Seq

Phage Immunoprecipitation-Sequencing (PhIP-Seq) allows for unbiased, proteome-wide autoantibody discovery across a variety of disease settings, with identification of disease-specific autoantigens providing new insight into previously poorly understood forms of immune dysregulation. Despite several successful implementations of PhIP-Seq for autoantigen discovery, including our previous work (Vazquez et al. 2020), current protocols are inherently difficult to scale to accommodate large cohorts of cases and importantly, healthy controls. Here, we develop and validate a high throughput extension of PhIP-seq in various etiologies of autoimmune and inflammatory diseases, including APS1, IPEX, RAG1/2 deficiency, Kawasaki Disease (KD), Multisystem Inflammatory Syndrome in Children (MIS-C), and finally, mild and severe forms of COVID19. We demonstrate that these scaled datasets enable machine-learning approaches that result in robust prediction of disease status, as well as the ability to detect both known and novel autoantigens, such as PDYN in APS1 patients, and intestinally expressed proteins BEST4 and BTNL8 in IPEX patients. Remarkably, BEST4 antibodies were also found in 2 patients with RAG1/2 deficiency, one of whom had very early onset IBD. Scaled PhIP-Seq examination of both MIS-C and KD demonstrated rare, overlapping antigens, including CGNL1, as well as several strongly enriched putative pneumonia-associated antigens in severe COVID19, including the endosomal protein EEA1. Together, scaled PhIP-Seq provides a valuable tool for broadly assessing both rare and common autoantigen overlap between autoimmune diseases of varying origins and etiologies.

immunology↗

bioRxiv2021

An AI-guided invariant signature places MIS-C with Kawasaki disease in a continuum of host immune responses

A significant surge in cases of multisystem inflammatory syndrome in children (MIS-C, also called Pediatric Inflammatory Multisystem Syndrome - PIMS) has been observed amidst the COVID-19 pandemic. MIS-C shares many clinical features with Kawasaki disease (KD), although clinical course and outcomes are divergent. We analyzed whole blood RNA sequences, serum cytokines, and formalin fixed heart tissues from these patients using a computational toolbox of two gene signatures, i.e., the 166-gene viral pandemic (ViP) signature, and its 20-gene severe (s)ViP subset that were developed in the context of SARS-CoV-2 infection and a 13-transcript signature previously demonstrated to be diagnostic for KD. Our analyses revealed that KD and MIS-C are on the same continuum of the host immune response as COVID-19. While both the pediatric syndromes converge upon an IL15/IL15RA-centric cytokine storm, suggestive of shared proximal pathways of immunopathogenesis, they diverge in other laboratory parameters and cardiac phenotypes. The ViP signatures also revealed unique targetable cytokine pathways in MIS-C, place MIS-C farther along in the spectrum in severity compared to KD and pinpoint key clinical (reduced cardiac function) and laboratory (thrombocytopenia and eosinopenia) parameters that can be useful to monitor severity.

immunology↗

bioRxiv2021

Kawasaki Disease patient stratification and pathway analysis based on host transcriptomic and proteomic profiles

The aetiology of Kawasaki Disease (KD), an acute inflammatory disorder of childhood, remains unknown despite various triggers of KD having been proposed. Host omic profiles offer insights into the host response to infection and inflammation, with the interrogation of multiple omic levels in parallel providing a more comprehensive picture. We used differential abundance analysis, pathway analysis, clustering and classification techniques to explore whether the host response in KD is more similar to the response to bacterial or viral infection at the transcriptomic and proteomic levels through comparison of omic profiles from children with KD to those with bacterial and viral infections. Pathways activated in patients with KD included those involved in anti-viral and anti-bacterial responses. Unsupervised clustering showed that the majority of KD patients clustered with bacterial patients on both omic levels, whilst application of diagnostic signatures specific for bacterial and viral infections revealed that many transcriptomic KD samples had low probabilities of having bacterial or viral infections, suggesting that KD may be triggered by a different process not typical of either common bacterial or viral infections. Clustering based on the transcriptomic and proteomic responses during KD revealed three clusters of KD patients on both omic levels, suggesting heterogeneity within the inflammatory response during KD. The observed heterogeneity may reflect differences in the host response to a common trigger, or variation dependent on different triggers of the condition.

bioinformatics↗

bioRxiv2020

CUBIC-Cloud: An Integrative Computational Framework Towards Community-Driven Whole-Mouse-Brain Mapping

Recent advancements in tissue clearing technologies have offered unparalleled opportunities for researchers to explore the whole mouse brain at cellular resolution. With the expansion of this experimental technique, however, a scalable and easy-to-use computational tool is in demand to effectively analyze and integrate whole-brain mapping datasets. To that end, here we present CUBIC-Cloud, a cloud-based framework to quantify, visualize and integrate whole mouse brain data. CUBIC-Cloud is a fully automated system where users can upload their whole-brain data, run analysis and publish the results. We demonstrate the generality of CUBIC-Cloud by a variety of applications. First, we investigated brain-wide distribution of PV, Sst, ChAT, Th and Iba1 expressing cells. Second, A{beta} plaque deposition in AD model mouse brains were quantified. Third, we reconstructed neuronal activity profile under LPS-induced inflammation by c-Fos immunostaining. Last, we show brain-wide connectivity mapping by pseudo-typed Rabies virus. Together, CUBIC-Cloud provides an integrative platform to advance scalable and collaborative whole-brain mapping.

neuroscience↗