Biology subjects

Iwamoto, H.

Publications and source records attributed to Iwamoto, H..

10 recordsLinked to original sources

bioRxiv2026

Androgen receptor determines skeletal muscle sexual dimorphism

The molecular and functional bases of sexual dimorphism in skeletal muscle remain poorly understood. The androgen receptor (AR) is a major regulator of sex-biased gene expression in muscle, but its genomic targets and associated coregulators in vivo are incompletely defined. Using ChIL-seq and an AirID-AR knock-in mouse, we mapped AR-bound genes and AR-associated proteins in skeletal muscle and identified histone deacetylase-linked corepressors. We further identified myosin binding protein H (Mybph) as a female-biased AR-repressed gene conserved in mouse and human muscle. Mybph loss disrupted sarcomeric organization and selectively delayed postinjury force recovery in female mice. These findings define an in vivo AR regulatory network and identify AR-dependent Mybph repression as a potential mechanism contributing to skeletal muscle sexual dimorphism.

molecular biology↗

bioRxiv2025

An atlas of microtubule lattice parameters regulated through ligand binding to the microtubule stabilizing sites.

Microtubules (MTs) are dynamic cytoskeletal polymers whose lattice architecture regulates force generation, nucleotide hydrolysis, and recognition by motor proteins and microtubule-associated proteins (MAPs). Microtubule-stabilizing agents (MSAs), including taxanes and laulimalide/peloruside-site ligands, suppress depolymerization by binding to defined lattice sites, yet stabilization is not structurally neutral. How ligand chemistry reshapes lattice organization and function remains unresolved. Here, we address three mechanistic questions. First, do distinct ligand classes induce defined lattice states? Using X-ray fiber diffraction, we show that MSAs selectively stabilize two preferred longitudinal conformations, a compact state ([~]4.06 nm monomer rise) and an expanded state ([~]4.17 nm), while modulating lateral organization reflected in shifts in mean MT radius. These axial spacings cluster around discrete values across chemotypes, indicating stabilization of pre-existing conformational minima rather than continuous distortion. Second, are these states interconvertible upon changes in ligand occupancy? Time-resolved diffraction reveals that longitudinal transitions occur within seconds of ligand addition even at substoichiometric occupancy; whereas, lateral equilibration proceeds slower, consistent with redistribution within heterogeneous protofilament organizations. Third, do such structural states alter nucleotide hydrolysis and motor/MAP behavior? Expanded lattices are associated with reduced apparent GTP hydrolysis rates under steady-state assembly conditions and altered kinesin motility, whereas compact lattices preferentially promote tau binding and distinct motor interaction profiles. Together, these findings establish longitudinal lattice conformation as a regulatory parameter and position microtubule-stabilizing agents as chemical tools that bias a dynamic structural landscape with predictable catalytic and transport consequences. Significance statementMicrotubules generate force and support intracellular transport through lattice geometries selectively recognized by motor proteins and microtubule-associated proteins. Microtubule-stabilizing drugs such as taxanes and epothilones are widely used in chemotherapy but can cause neurotoxicity, likely because stabilization alters lattice architecture rather than simply preventing depolymerization. Here, we show that stabilizing ligands bias microtubules between two preferred longitudinal conformations, compact and expanded, that can switch within seconds of binding, while lateral organization adjusts slower. These structural states differentially regulate GTP hydrolysis and recognition by tau and kinesin, linking lattice geometry to catalytic and transport functions. By establishing lattice conformation as a tunable regulatory parameter, this work provides a framework for interpreting drug effects and designing structure-selective stabilizers with improved therapeutic profiles.

biophysics↗

bioRxiv2025

Peripheral Inflammation Limits Serotonin Neuron Signaling Capacity via Serotonergic IL-1R1 to Reduce Neuronal Excitability and Enhance Serotonin Clearance

Neurobehavioral disorders, ranging from depression to schizophrenia, have been found to display immune system alterations. The high incidence of comorbidity of these disorders, particularly depression, with chronic inflammatory conditions suggests shared mechanisms contributing to the manifestation of these conditions. We have previously shown that peripheral modulation of the innate immune system in mice rapidly triggers enhanced serotonin (5-HT) clearance in vivo associated with increased anxiety- and despair-like behaviors that can be suppressed by serotonergic elimination of p38 MAPK. Forebrain-projecting 5-HT synthesizing neurons of the dorsal raphe nucleus (DRN5-HT) play a key role in regulating behaviors related to mood and anxiety and whose perturbations are observed in multiple affective disorders. Here we identify molecular and circuit-level mechanisms that can translate peripheral innate immune system activation into changes in 5-HT signaling capacity. Using whole cell patch clamp recordings from acute midbrain slices, we demonstrate that the proinflammatory cytokine, IL-1{beta}, acts cell autonomously through its receptor, IL-1R1, via the p38 MAPK signaling pathway to rapidly inhibit firing of DRN5-HT neurons. In the dorsal hippocampus, we found that as with acute, peripheral lipopolysaccharide (LPS) administration, local injections of IL-1{beta} rapidly enhance 5-HT clearance as assessed by in vivo chronoamperometry. Like IL-1{beta}, TNF also acts via a serotonergic p38 MAPK dependent pathway to reduce excitability of DRN5-HT neurons. Immunocytochemical studies reveal that, IL-1R1, is nonuniformly expressed by DRN5-HT neurons and is required for LPS-induced inhibition of these cells as detected by cFos activation, with sex-dependent patterns evident. Moreover, we detected both DRN5-HT IL-1R1-dependent and -independent LPS-mediated changes in cFos changes in forbrain projection areas. Our findings support a growing appreciation that serotoninergic neurons contribute to changes in CNS physiology and behavior following peripheral immune activation. More specifically, our studies attest to a functional role of serotonergic IL-1R1 in mediating IL-1{beta} following peripheral innate immune activation, effects likely to arise both from changes in diminished 5-HT neuron excitability and elevated 5-HT clearance.

neuroscience↗

bioRxiv2024

Restoration of 3-D structure of insect flight muscle from a rotationally averaged 2-D X-ray diffraction pattern

The contractile machinery of muscle, especially that of skeletal muscle, has a very regular array of contractile protein filaments, and gives rise to a complex and informative diffraction pattern when irradiated with X-rays. However, analyzing these diffraction patterns is often challenging because: (1) only rotationally averaged diffraction patterns can be obtained, resulting in a substantial loss of information, and (2) the contractile machinery contains two different sets of protein filaments (actin and myosin) with different helical symmetries. The reflections originating from them often overlap. These problems may be solved if the real-space 3-D structure of the contractile machinery is directly calculated from the diffraction pattern. Here we demonstrate that, by using the conventional phase-retrieval algorithm (hybrid input-output), the real-space 3-D structure of the contractile machinery can be effectively restored from a single rotationally averaged 2-D diffraction pattern. In this calculation, we used an in-silico model of insect flight muscle, which is known for its highly regular structure. We also extended this technique to an experimentally recorded muscle diffraction pattern.

biophysics↗

bioRxiv2024

Dysregulated Skeletal Muscle Myosin Super-relaxation in Type II, but Not Type I, Diabetes Mellitus

Disrupted energy balance is critical for the onset and development of Type II diabetes. The exact underlying metabolic mechanisms remain incomplete but skeletal muscle is thought to play an important pathogenic role. As the super-relaxed state of its most abundant protein, myosin, regulates cellular energetics, here, we aimed to investigate whether it is altered in patients with type II diabetes. For that, we used vastus lateralis biopsy specimens (obtained from patients with type II diabetes and matched controls) and run a combination of structural and functional assays consisting of loaded Mant-ATP chase experiments, X-ray diffraction and LC-MS/MS proteomics in isolated muscle fibres. Our studies revealed a greater muscle myosin super-relaxation and decreased cellular ATP demand in patients than controls. Subsequent proteomic analyses indicated that these (mal)adaptations likely originated from remodeled sarcomeric proteins and greater myosin glycation levels in patients than controls. Overall, our findings emphasize a complex molecular dysregulation of myosin super-relaxed state and energy consumption in type II diabetes. Ultimately, pharmacological targeting of myosin could benefit skeletal muscle and whole-body metabolic health through the enhancement of ATP consumption. Significance StatementMyosin super-relaxation, essential for the regulation of skeletal muscle metabolic rate, is disrupted in type II diabetes due to protein hyper-glycation. As a consequence, myosin ATP demand is significantly lowered. Overall, our findings provide a strong rationale for the use of activators of myosin ATPase to enhance basal energy expenditure in type II diabetes.

cell biology↗

bioRxiv2023

Remodelling of Skeletal Muscle Myosin Metabolic States in Hibernating Mammals

Hibernation is a period of metabolic suppression utilized by many small and large mammal species to survive during winter periods. As the underlying cellular and molecular mechanisms remain incompletely understood, our study aimed to determine whether skeletal muscle myosin and its metabolic efficiency undergo alterations during hibernation to optimize energy utilization. We isolated muscle fibers from small hibernators, Ictidomys tridecemlineatus and Eliomys quercinus and larger hibernators, Ursus arctos and Ursus americanus. We then conducted loaded Mant-ATP chase experiments alongside X-ray diffraction to measure resting myosin dynamics and its ATP demand. In parallel, we performed multiple proteomics analyses. Our results showed a preservation of myosin structure in U. arctos and U. americanus during hibernation, whilst in I. tridecemlineatus and E. quercinus, changes in myosin metabolic states during torpor unexpectedly led to higher levels in energy expenditure of type II, fast-twitch muscle fibers at ambient lab temperatures (20{degrees}C). Upon repeating loaded Mant-ATP chase experiments at 8{degrees}C (near the body temperature of torpid animals), we found that myosin ATP consumption in type II muscle fibers was reduced by 77-107% during torpor compared to active periods. Additionally, we observed Myh2 hyper-phosphorylation during torpor in I. tridecemilineatus, which was predicted to stabilize the myosin molecule. This may act as a potential molecular mechanism mitigating myosin-associated increases in skeletal muscle energy expenditure during periods of torpor in response to cold exposure. Altogether, we demonstrate that resting myosin is altered in hibernating mammals, contributing to significant changes to the ATP consumption of skeletal muscle. Additionally, we observe that it is further altered in response to cold exposure and highlight myosin as a potentially contributor to skeletal muscle non-shivering thermogenesis.

physiology↗

bioRxiv2023

Human light meromyosin mutations linked to skeletal myopathies disrupt the coiled coil structure and myosin head sequestration

Myosin heavy chains encoded by MYH7 and MYH2 are among the most abundant proteins in human skeletal muscle. After decades of intense research using a wide range of biophysical and biological approaches, their functions have begun to be elucidated. Despite this, it remains unclear how mutations in these genes and resultant proteins disrupt myosin structure and function, inducing pathological states and skeletal myopathies termed myosinopathies. Here, we have analysed the effects of several common MYH7 and MYH2 mutations located in light meromyosin (LMM) using a broad range of approaches. We determined the secondary structure and filament forming capabilities of expressed and purified LMM constructs in vitro, performed in-silico modelling of LMM constructs, and evaluated the incorporation of eGFP-myosin heavy chain constructs into sarcomeres in cultured myotubes. Using muscle biopsies from patients, we applied Mant-ATP chase protocols to estimate the proportion of myosin heads that were super-relaxed, X-ray diffraction measurements to estimate myosin head order and myofibre mechanics to investigate contractile function. We found that human MYH7 and MYH2 LMM mutations commonly disrupt myosin coiled-coil structure and packing of filaments in vitro; decrease the myosin super-relaxed state in vivo and increase the basal myosin ATP consumption; but are not associated with myofibre contractile deficits. Altogether, these findings indicate that the structural remodelling resulting from LMM mutations induces a pathogenic state in which formation of shutdown heads is impaired, thus increasing myosin head ATP demand in the filaments, rather than affecting contractility. These key findings will help in the design of future therapies for myosinopathies.

physiology↗

bioRxiv2023

Predominant myosin super-relaxed state in canine myocardium with naturally occurring dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a naturally occurring heart failure condition in humans and dogs, notably characterized by a reduced contractility and ejection fraction. As the identification of its underlying cellular and molecular mechanisms remain incomplete, the aim of the present study was to assess whether the molecular motor myosin and its known relaxed conformational states are altered in DCM. For that, we dissected and skinned thin cardiac strips from left ventricle obtained from six DCM Doberman Pinschers and six non-failing controls (NF). We then used a combination of Mant-ATP chase experiments and X-ray diffraction to assess both energetic and structural changes of myosin. Using the Mant-ATP chase protocol, we observed that in DCM dogs, the amount of myosin molecules in the ATP-conserving conformational state also known as super-relaxed (SRX), is significantly increased when compared with NF dogs. This alteration can be rescued by applying EMD-57033, a small molecule activating myosin. Conversely, with X-ray diffraction, we found that in DCM dogs, there is a higher proportion of myosin heads in the vicinity of actin when compared with NF dogs (1,0 to 1,1 intensity ratio). Hence, we observed an uncoupling between energetic (Mant-ATP chase) and structural (X-ray diffraction) data. Taken together, these results may indicate that in the heart of Doberman Pinschers with DCM, myosin molecules are potentially stuck in a non-sequestered but ATP-conserving SRX state, that can be counterbalanced by EMD-57033 demonstrating the potential for a myosin-centered pharmacological treatment of DCM. New & noteworthyThe key finding of the present study is that, in left ventricles of dogs with a naturally occurring dilated cardiomyopathy, relaxed myosin molecules favor a non-sequestered super-relaxed state potentially impairing sarcomeric contractility. This alteration is rescuable by applying a small molecule activating myosin known as EMD-57033.

pathology↗

bioRxiv2023

Serotonin in the orbitofrontal cortex enhances cognitive flexibility

Cognitive flexibility is a brains ability to switch between different rules or action plans depending on the context. However, cellular level understanding of cognitive flexibility have been largely unexplored. We probed a specific serotonergic pathway from dorsal raphe nuclei (DRN) to the orbitofrontal cortex (OFC) while animals are performing reversal learning task. We found that serotonin release from DRN to the OFC promotes reversal learning. A long-range connection between these two brain regions was confirmed anatomically and functionally. We further show that spatiotemporally precise serotonergic action directly enhances the excitability of OFC neurons and offers enhanced spike probability of OFC network. Serotonergic action facilitated the induction of synaptic plasticity by enhancing Ca2+ influx at dendritic spines in the OFC. Thus, our findings suggest that a key signature of flexibility is the formation of choice specific ensembles via serotonin-dependent synaptic plasticity.

neuroscience↗

bioRxiv2021

The 3-D structure of fibrous material is fully restorable from its X-ray diffraction pattern

X-ray fiber diffraction is potentially a powerful technique to study the structure of fibrous materials, such as DNA and synthetic polymers. However, only rotationally averaged diffraction patterns can be recorded, and it is difficult to correctly interpret them without the knowledge of esoteric diffraction theories. Here we demonstrate that, in principle, the non-rotationally averaged 3-D structure of the material can be restored from its fiber diffraction pattern. The method is a simple puzzle-solving process, and in ideal cases, it does not require any prior knowledge about the structure, such as helical symmetry. We believe that the proposed method has a potential to transform the fiber diffraction to a 3-D imaging technique, and will be useful for a wide field of life and materials sciences.

biophysics↗