Biology subjects

Ogura, T.

Publications and source records attributed to Ogura, T..

8 recordsLinked to original sources

bioRxiv2024

Structural and Dynamic Insights into the Biased Signaling Mechanism of the Human Kappa Opioid Receptor

The {kappa}-opioid receptor (KOR) is a member of the G protein-coupled receptor (GPCR) family, responsible for modulating cellular responses through transducers such as G proteins and arrestins. G protein-biased KOR agonists hold promise due to their potential to mitigate side effects such as drug aversion and sedation while preserving analgesic and antipruritic effects. Here, we shed light on the structural dynamics of the human KOR-Gi signaling complex bound with either nalfurafine (a G-protein-biased agonist) or U-50,488H (a balanced agonist) using cryo-electron microscopy (cryo-EM). Cryo-EM structures of the KOR-Gi signaling complexes identify the ligand binding mode in the activated state. Vibrational spectroscopy analysis reveals changes in the ligand-binding pocket upon binding to these ligands. Cell-based mutant experiments pinpoint four amino acids (K2275.40, C2866.47, H2916.52, and Y3127.34; Ballesteros-Weinstein numbering is shown in superscript) that play crucial roles in arrestin recruitment. Among these four amino acids, H2916.52 and Y3127.34 are also implicated in G-protein coupling. Our findings pave the way for targeting specific residues in the KOR ligand-binding pocket to enhance KOR-mediated therapeutic effects while mitigating unwanted side effects.

biophysics↗

bioRxiv2023

Protective effects of Mkl1/2 against lipodystrophy and muscle atrophy via PI3K/AKT-independent FoxO repression

FoxO transcription factors are involved in the pathogenesis of lipodystrophy and muscle atrophy. FoxO proteins are phosphorylated and inactivated by PI3K/AKT signaling; however, little is known about FoxO repressors other than this pathway. Our study showed that the Srf cofactors Mkl1 and Mkl2 directly repressed FoxO transcriptional activity, independent of the PI3K/AKT pathway. Loss of Mkl1/2 led to the overactivation of FoxO, which impaired the maintenance of both white adipose tissue (WAT) and skeletal muscle. In Mkl2-deficient preadipocytes, Ppar{gamma} was suppressed and white adipogenesis was severely impaired. In myotubes, Mkl1 and Mkl2 suppressed the expression of atrophy-related genes (atrogenes) induced by FoxO. Mkl1 expression was reduced at the onset of muscle atrophy in vivo, and exogenous supplementation of skeletal muscle Mkl1 suppressed atrogene expression and induced muscle hypertrophy. Finally, both Mkl2 nuclear localization and Mkl1/2 expression were upregulated by exercise, suggesting that Mkl1/2 is involved in the inhibitory effect of exercise on muscle atrophy. These findings indicate that Mkl1/2 act as PI3K/AKT signaling-independent repressors of FoxO and are essential for adipose and muscle tissue homeostasis.

cell biology↗

bioRxiv2023

Latent inter-organ mechanism of idiopathic pulmonary fibrosis unveiled by a generative computational approach

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive disease characterized by complex lung pathogenesis affecting approximately three million people worldwide. While the molecular and cellular details of the IPF mechanism is emerging, our current understanding is centered around the lung itself. On the other hand, many human diseases are the products of complex multi-organ interactions. Hence, we postulate that a dysfunctional crosstalk of the lung with other organs plays a causative role in the onset, progression and/or complications of IPF. In this study, we employed a generative computational approach to identify such inter-organ mechanism of IPF. The approach works as follows: 1) To find unexpected relatedness of IPF to other diseases of non-lung organs and to identify molecular features that define such relatedness, 2) To identify differentially expressed genes between the lung tissues of IPF vs. those of non-IPF pulmonary disease patients, 3) To detect ligand-receptor relationships across multiple organs and their upstream and downstream signaling pathways in 1) and 2), 4) To generate a map of the inter-organ IPF mechanism with the molecular and cellular resolution. This approach found unexpected molecular relatedness of IPF to neoplasm, diabetes, Alzheimers disease, obesity, atherosclerosis, and arteriosclerosis. Furthermore, as a potential mechanism underlying this relatedness, we uncovered a putative molecular crosstalk system across the lung and the liver. In this inter-organ system, a secreted protein, kininogen 1, from hepatocytes in the liver interacts with its receptor, bradykinin receptor B1 in the lung. This ligand-receptor interaction across the liver and the lung leads to the activation of calmodulin pathways in the lung, leading to the activation of interleukin 6 and phosphoenolpyruvate carboxykinase 1 pathway across these organs. Furthermore, we retrospectively identified several pre-clinical and clinical evidence supporting this inter-organ mechanism of IPF. In conclusion, such feedforward and feedback system across the lung and the liver provides a unique opportunity for the development of the treatment and/or diagnosis of IPF. Furthermore, the result illustrates a generative computational framework for machine-mediated synthesis of mechanisms that facilitates and complements the traditional experimental approaches in biomedical sciences.

pathology↗

bioRxiv2022

Isotope-based visualization of element distribution in phloem provides functional evidence for the operation of SOS1 Na+/H+ exchangers in mature zones of Arabidopsis root

In the phloem, various solutes such as photosynthates, mineral nutrients, or toxic elements move from the source organs to sinks. One of them is the root. While this process may mediate significant quantities of solutes delivered to the root, it is unclear how the solutes are distributed along the root axis. To elucidate the distributing pattern of solutes, we visually analyzed the movement of solutes in Arabidopsis roots using a radioisotope imaging technique. The distribution patterns fell into four different groups: (1) root tip accumulation for 14C-photosynthates, 28Mg2+, 32PO43-, and 35SO42-; (2) homogenous distribution along the root axis for 42K+ and 137Cs+; (3) no detectable accumulation in the root for 45Ca2+ and 59Fe2+; and (4) transient accumulation in the root for 22Na+. The latter phenomenon was explored in more detail using Arabidopsis knockout mutants lacking functional sodium efflux transporter SOS1 (operating as Na+/H+ exchanger). By utilizing a non-invasive microelectrode MIFE ion flux measuring technique, we found that Na+ efflux was active in the mature root zone of wild-type Arabidopsis plants but not in root apex as initially thought, and that in sos1 mutants lacking functional Na+/H+ exchangers, shoot-derived 22Na+ remained in the root. These findings challenge the notion that Na+ exclusion via SOS1 is confined to the root apex and demonstrates the power of combining the radioisotope imaging technique and the MIFE ion flux measuring technique to study the kinetics of ion transport in the root and root-to-shoot communication.

plant biology↗

bioRxiv2021

Distinct roles and actions of PDI family enzymes in catalysis of nascent-chain disulfide formation

The mammalian endoplasmic reticulum (ER) harbors more than 20 members of the protein disulfide isomerase (PDI) family that act to maintain proteostasis. Herein, we developed an in vitro system for directly monitoring PDI- or ERp46-catalyzed disulfide bond formation in ribosome-associated nascent chains (RNC) of human serum albumin. The results indicated that ERp46 more efficiently introduced disulfide bonds into nascent chains with short segments exposed outside the ribosome exit site than PDI. Single-molecule analysis by high-speed atomic force microscopy further revealed that PDI binds nascent chains persistently, forming a stable face-to-face homodimer, whereas ERp46 binds for a shorter time in monomeric form, indicating their different mechanisms for substrate recognition and disulfide bond introduction. Similarly to ERp46, a PDI mutant with an occluded substrate-binding pocket displayed shorter-time RNC binding and higher efficiency in disulfide introduction than wild-type PDI. Altogether, ERp46 serves as a more potent disulfide introducer especially during the early stages of translation, whereas PDI can catalyze disulfide formation in RNC when longer nascent chains emerge out from ribosome.

biochemistry↗

bioRxiv2020

Hierarchical Model for the Role of J-Domain Proteins in Distinct Cellular Functions

In Escherichia coli, the major bacterial Hsp70 system consists of DnaK, three J-domain proteins (JDPs: DnaJ, CbpA, and DjlA), and one nucleotide exchange factor (NEF: GrpE). JDPs determine substrate specificity for the Hsp70 system; however, knowledge on their specific role in bacterial cellular functions is limited. In this study, we demonstrated the role of JDPs in bacterial survival during heat stress and the DnaK-regulated formation of curli--extracellular amyloid fibers involved in E. coli biofilm formation. Genetic analysis with a complete set of JDP-null mutant strains demonstrated that only DnaJ is essential for survival at high temperature, while DnaJ and CbpA are indispensable in DnaK regulation of curli production. Additionally, we found that DnaJ and CbpA are involved in the expression of the master regulator CsgD through the folding of MlrA; this keeps CsgA in a translocation-competent state by preventing its aggregation in the cytoplasm. Our findings support a hierarchical model wherein the role of JDPs in the Hsp70 system differs according to individual cellular functions.

molecular biology↗

bioRxiv2019

EXOCYST70A3 controls root system depth in Arabidopsis via the dynamic modulation of auxin transport

Root system architecture (RSA), the distribution of roots in soil, plays a major role in plant survival. RSA is shaped by multiple developmental processes that are largely governed by the phytohormone auxin, suggesting that auxin regulates responses of roots that are important for local adaptation. However, auxin has a central role in numerous processes and it is unclear which molecular mechanisms contribute to the variation in RSA for environmental adaptation. Using natural variation in Arabidopsis, we identify EXOCYST70A3 as a modulator of the auxin system that causes variation in RSA by acting on PIN4 protein distribution. Allelic variation and genetic perturbation of EXOCYST70A3 leads to alteration of root gravitropic responses, resulting in a different RSA depth profile and drought resistance. Overall our findings suggest that the local modulation of the pleiotropic auxin pathway can gives rise to distinct root system architectures that can be adaptive in specific environments.

plant biology↗

bioRxiv2018

Pirfenidone attenuates lung fibrotic fibroblast-mediated fibrotic responses to transforming growth factor-β1

Pirfenidone, an antifibrotic agent used for treatment of idiopathic pulmonary fibrosis (IPF), functions by inhibiting myofibroblast differentiation, which is involved in transforming growth factor (TGF)-{beta}1-induced IPF pathogenesis. However, unlike normal lung fibroblasts, the relationship between pirfenidone responses of TGF-{beta}1-induced human fibrotic lung fibroblasts and lung fibrosis is unknown. Here, we investigated the effect of pirfenidone on the functions of two new targets, collagen triple helix repeat containing protein 1 (CTHRC1) and four-and-a-half LIM domain protein 2 (FHL2), which included fibroblast activity, collagen gel contraction, and migration toward fibronectin. Compared to control lung fibroblasts, pirfenidone restored TGF-{beta}1-stimulated fibroblast-mediated collagen gel contraction, migration, and CTHRC1 release in lung fibrotic fibroblasts. Furthermore, pirfenidone attenuated TGF-{beta}1- and CTHRC1-induced fibroblast activity, bone morphogenic protein-4/Gremlin1 upregulation, and -smooth muscle actin, fibronectin, and FHL2 downregulation, similar to that observed post-CTHRC1 inhibition. In contrast, FHL2 inhibition suppressed migration and fibronectin expression but did not downregulate CTHRC1. Overall, pirfenidone suppressed fibrotic fibroblast-mediated fibrotic processes via inverse regulation of CTHRC1-induced lung fibroblast activity. Thus, CTHRC1 can be used for predicting pirfenidone response and developing new therapeutic target for lung fibrosis.\n\nSummary statementPirfenidone suppressed TGF-{beta}1-mediated fibrotic processes in fibrotic lung fibroblasts by attenuating CTHRC1 expression, suggesting that CTHRC1 may be a novel therapeutic target for treating patients with lung fibrosis.

cell biology↗