Biology subjects

Ozawa, K.

Publications and source records attributed to Ozawa, K..

8 recordsLinked to original sources

bioRxiv2026

Neuropeptide T2A-GAL4 knock-ins illustrate practical considerations for interpreting GAL4 reporter patterns in Drosophila

Transgenic GAL4 lines are widely used in Drosophila to label and manipulate specific cell types, but the resulting reporter expression patterns do not always represent ongoing driver activity. We generated a collection of neuropeptide T2A-GAL4 knock-ins and characterized their expression by native-fluorescence imaging using UAS-CD8::GFP and a Brp-miRFP680 knock-in, eliminating the need for immunostaining. During characterization of these lines, we observed cases in which adult GFP patterns did not match the on-going GAL4 activity, as well as cases in which GAL4 produced adverse cellular effects. Using temperature-sensitive GAL80, we found that adult GFP expression in certain neurons driven by CCHa2-T2A-GAL4 and Dsk-T2A-GAL4 depended on GAL4 activity before eclosion and was not reproduced by adult-only induction. Several GAL4 lines also showed dominant developmental phenotypes. Heterozygous Burs-T2A-GAL4 animals, for example, displayed wing-expansion defects that were fully suppressed by GAL80. Severe wing defects were not accompanied by a reduction in the number of reporter-labeled neurons before eclosion, but the morphology of these neurons were affected. These observations illustrate how developmental driver history and GAL4-associated cellular effects can influence reporter patterns in the adult nervous system.

genetics↗

bioRxiv2026

YY1 Binding Motif at Upstream of Rep/Cap Increases AAV Yield and Full Capsids

Adeno-associated virus (AAV) vectors are widely used in gene therapy, whereas low manufacturing efficiency and a large proportion of empty capsids are major obstacles. This study focused on the Yin Yang 1 (YY1) binding motif (YY1-motif) and investigated the effect of its presence or insertion upstream of the Replicase (Rep)/Capsid (Cap) gene on AAV vector production. We found that the YY1-motif incidentally presented in a Rep/Cap plasmid was associated with high vector production. We then designed several modified Rep/Cap (RC2) constructs. The YY1-motif insertion in front of Rep/Cap gene increased vector yield in a repeat-number-dependent manner, and similar effects were not observed with other promoters insertion. Furthermore, the insertion of the YY1-motif reduced the amount of Cap protein per the same amount of full particle in supernatants on multiple serotypes, indicating the improvement in the empty/full capsid ratio. The YY1-motif insertion did not affect the AAV vector infectivity. These results denote that the YY1-motif has a universal regulatory function that optimizes the Rep/Cap expression balance, and simultaneously improves the production efficiency and full particle formation of AAV vectors. This finding could contribute to the development of highly efficient and high-quality AAV manufacturing processes.

microbiology↗

bioRxiv2025

Human-specific fast synaptic kinetics enable rapid detection of predictive features

Humans can discern the underlying principles in complex environments, but the cellular basis of this ability is unclear. By comparing layer 2/3 cortical pyramidal neurons in five mammals (mouse, rat, marmoset, macaque and human), we found that human neurons exhibited distinctively faster kinetics of excitatory synaptic transmissions. In a computational model, neurons with human-type synapses detected hidden patterns more quickly in noise, and this superiority was only evident in unsupervised learning. Tracking synaptic weights revealed that human synapses were potentiated sharply and persistently by the patterns, while macaque-type synapses were broadly potentiated, which were easily eroded. These data suggest that the human-specific acceleration of synaptic kinetics enables the rapid extraction of latent structures in complex environments, providing a cellular basis for our efficient cognitive abilities.

neuroscience↗

bioRxiv2025

AutoLead: An LLM-Guided Bayesian Optimization Framework for Multi-Objective Lead Optimization

The process of lead optimization in drug discovery is a complex, multi-objective challenge that remains a major bottleneck in the development of new therapeutics. Traditional approaches often struggle to efficiently explore the vast chemical space while simultaneously optimizing multiple, and sometimes conflicting, molecular properties. In this work, we present AutoLead, a novel framework that integrates Large Language Models (LLMs) with multi-objective Bayesian optimization to tackle this challenge. By leveraging the chemical reasoning capabilities of LLMs, AutoLead effectively guides the search for novel drug-like molecules that satisfy multiple objectives. We evaluate our approach on two molecule optimization tasks, achieving state-of-the-art results. Furthermore, we introduce a new benchmark dataset designed around a more realistic lead optimization scenario, where the task is to modify compounds that violate Lipinskis Rule of Five to simultaneously meet all criteria and improve their QED score. Through extensive experiments and a detailed case study, we demonstrate the potential of combining LLMs with black-box optimization techniques for more efficient and practical drug discovery.

bioinformatics↗

bioRxiv2024

Fetal Kidney Transplantation for In Utero Fetuses

Potter sequence, characterized by bilateral renal agenesis, oligohydramnios, and consequent pulmonary hypoplasia, presents a significant challenge in the management of affected neonates. Due to their prematurity and associated abdominal complications, these infants often fail to reach a stage where dialysis can be safely initiated and sustained, leading to an exceedingly high mortality rate. Therefore, there is hopeful anticipation that interventions serving as a bridge to achieve a state where dialysis can be safely performed will markedly improve life expectancy. We have developed a unique approach of "transplantation of fetal kidneys from a different species during the fetal period" as a bridge therapy until stable dialysis therapy can be implemented. This is a new concept of fetal therapy, targeting the fetus in utero and utilizing fetal kidneys of an appropriate size for transplantation. In this study, we first validated the approach using allogeneic transplantation. Fetal kidneys with bladders from GFP-expressing rats (gestational age 14.0-16.5 days) were transplanted subcutaneously into allogeneic rat fetuses in utero (gestational age 18.0-18.5 days) using a special needle transuterinally, and live pups were successfully obtained. The transplanted fetal kidneys with bladders were confirmed to have urine production capability. By periodic aspiration of the subcutaneous urinary cyst after birth, urine produced by the transplanted fetal kidney was successfully drained outside the body for an extended period (up to 150 days). Biochemical tests confirmed the solute removal capacity of the transplanted fetal kidney. Furthermore, despite allogeneic transplantation, long-term urine production was sustained without the use of immunosuppressants, confirming that organ transplantation into fetuses is associated with lower rejection compared to adult transplantation. Next, xenotransplantation was performed. When GFP-expressing mouse fetal kidneys (gestational age 13.0-13.5 days) were transplanted into rat fetuses in utero, maturation of renal tissue structures was confirmed even in the interspecies setting.

bioengineering↗

bioRxiv2023

Dynamics of motor direction representation in the primate premotor and primary motor cortices during sensorimotor learning

Sensorimotor learning requires reorganization of neuronal activity in the premotor cortex (PM) and primary motor cortex (M1). However, how PM- and M1-specific reorganization occurs in primates remains unclear. We conducted calcium imaging of these areas in common marmosets while they learned a two-target reaching (pull/push) task. Throughout learning, the dorsorostral PM (PMdr) showed peak activity earlier than the dorsocaudal PM (PMdc) and M1. PMdr showed decreased representation of newly introduced (push) movement, whereas PMdc and M1 maintained high representation. Many task-related neurons in PMdc and M1 exhibited a strong preference to either movement direction. PMdc neurons dynamically switched their preferred direction, whereas M1 neurons stably retained their preferred direction. Differences in preferred direction between adjacent neurons in PMdc increased during learning. These results suggest that in primate sensorimotor learning, dynamic motor representation in PMdc converts the cognitive sensorimotor signals of PMdr to stable and specific motor representation of M1.

neuroscience↗

bioRxiv2022

Effects of Simvastatin on RBL-2H3 cell degranulation.

Hypercholesterolemia is a major complication of arteriosclerosis. Mast cells in arteriosclerosis plaques induce inflammatory reactions, and promote arterial sclerosis. In this study, we evaluated the pharmacological effects of simvastatin (SV)-3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitors on the degranulation of rat basophilic leukemia (RBL)-2H3 cells, which are commonly used as mast cell models. SV significantly decreased degranulation induced by three types of stimulation: antigen antibody reaction (Ag-Ab), thapsigargin (Tg) serosal endoplasmic reticulum calcium ATPase (SERCA) inhibitor, and A23187 calcium ionophore. SV had a stronger inhibitory effect on degranulation induced by Ag-Ab stimulation than the other two stimulations. However, SV had no effect on increases of intracellular Ca2+ concentrations. Mevalonate or geranylgeraniol co-treatment with SV completely prevented the inhibitory effect of SV on degranulation induced by these stimulations. Immunoblotting results showed that SV inhibited protein kinase C (PKC) delta translocation that was induced by Ag-Ab but not by Tg or A23187. SV induced a reduction in active Rac1, and actin filament rearrangements. In conclusion, SV inhibits RBL-2H3 cell degranulation by inhibiting downstream signaling pathways, including the sequential degranulation pathway. These inhibitory effects were completely reversed by the addition of geranylgeraniol and might be induced by changes in the translocation of the small GTPase families Rab and Ras and Rho, which are related to vesicular transport and PKC delta activation, respectively. These changes are caused by the inhibition of HMG-CoA reductase by SV following the synthesis of farnesyl and geranylgeranyl pyrophosphates, which play important roles in the activation of small GTPases.

pharmacology and toxicology↗

bioRxiv2019

Transient astrocytic Gq signaling underlies remote memory enhancement

Astrocytes elicit transient Ca2+ elevations induced by G protein-coupled receptors (GPCRs), yet their role in vivo remains unknown. To address this, transgenic mice with astrocytic expression of the optogenetic Gq-type GPCR, Opto1AR, were established, in which transient Ca2+ elevations similar to those in wild type mice were induced by brief blue light illumination. Activation of cortical astrocytes resulted in an adenosine A1 receptor-dependent inhibition of neuronal activity. Moreover, sensory stimulation with astrocytic activation induced long-term depression of sensory evoked response. At the behavioral level, repeated astrocytic activation in the anterior cortex gradually affected novel open field exploratory behavior, and remote memory was enhanced in a novel object recognition task. These effects were blocked by A1 receptor antagonism. On the other hand, compelling evidence for astrocytic Ca2+-induced diameter changes of arteries was not observed. Together, we demonstrate that GPCR-triggered Ca2+ elevation in cortical astrocytes has causal impacts on neuronal activity and behavior.

neuroscience↗