基部陸上植物ゼニゴケのRAB GTPaseの研究

学位の種別: 課程博士審査委員会委員 : （主査）東京大学教授 塚谷 裕一, 東京大学教授 武田 洋幸, 東京大学教授 中野 明彦, 東京大学准教授 野﨑 久義, 基礎生物学研究所教授 上田 貴志University of Tokyo(東京大学

Harnessing Deep Learning to Analyze Cryptic Morphological Variability of Marchantia polymorpha

Characterizing phenotypes is a fundamental aspect of biological sciences, although it can be challenging due to various factors. For instance, the liverwort Marchantia polymorpha is a model system for plant biology and exhibits morphological variability, making it difficult to identify and quantify distinct phenotypic features using objective measures. To address this issue, we utilized a deep-learning-based image classifier that can handle plant images directly without manual extraction of phenotypic features and analyzed pictures of M. polymorpha. This dioicous plant species exhibits morphological differences between male and female wild accessions at an early stage of gemmaling growth, although it remains elusive whether the differences are attributable to sex chromosomes. To isolate the effects of sex chromosomes from autosomal polymorphisms, we established a male and female set of recombinant inbred lines (RILs) from a set of male and female wild accessions. We then trained deep learning models to classify the sexes of the RILs and the wild accessions. Our results showed that the trained classifiers accurately classified male and female gemmalings of wild accessions in the first week of growth, confirming the intuition of researchers in a reproducible and objective manner. In contrast, the RILs were less distinguishable, indicating that the differences between the parental wild accessions arose from autosomal variations. Furthermore, we validated our trained models by an ‘eXplainable AI’ technique that highlights image regions relevant to the classification. Our findings demonstrate that the classifier-based approach provides a powerful tool for analyzing plant species that lack standardized phenotyping metrics

Insights into Land Plant Evolution Garnered from the Marchantia polymorpha Genome.

The evolution of land flora transformed the terrestrial environment. Land plants evolved from an ancestral charophycean alga from which they inherited developmental, biochemical, and cell biological attributes. Additional biochemical and physiological adaptations to land, and a life cycle with an alternation between multicellular haploid and diploid generations that facilitated efficient dispersal of desiccation tolerant spores, evolved in the ancestral land plant. We analyzed the genome of the liverwort Marchantia polymorpha, a member of a basal land plant lineage. Relative to charophycean algae, land plant genomes are characterized by genes encoding novel biochemical pathways, new phytohormone signaling pathways (notably auxin), expanded repertoires of signaling pathways, and increased diversity in some transcription factor families. Compared with other sequenced land plants, M. polymorpha exhibits low genetic redundancy in most regulatory pathways, with this portion of its genome resembling that predicted for the ancestral land plant. PAPERCLIP

Comprehensive and quantitative analysis of intracellular structure polarization at the apical–basal axis in elongating Arabidopsis zygotes

Abstract A comprehensive and quantitative evaluation of multiple intracellular structures or proteins is a promising approach to provide a deeper understanding of and new insights into cellular polarity. In this study, we developed an image analysis pipeline to obtain intensity profiles of fluorescent probes along the apical–basal axis in elongating Arabidopsis thaliana zygotes based on two-photon live-cell imaging data. This technique showed the intracellular distribution of actin filaments, mitochondria, microtubules, and vacuolar membranes along the apical–basal axis in elongating zygotes from the onset of cell elongation to just before asymmetric cell division. Hierarchical cluster analysis of the quantitative data on intracellular distribution revealed that the zygote may be compartmentalized into two parts, with a boundary located 43.6% from the cell tip, immediately after fertilization. To explore the biological significance of this compartmentalization, we examined the positions of the asymmetric cell divisions from the dataset used in this distribution analysis. We found that the cell division plane was reproducibly inserted 20.5% from the cell tip. This position corresponded well with the midpoint of the compartmentalized apical region, suggesting a potential relationship between the zygote compartmentalization, which begins with cell elongation, and the position of the asymmetric cell division

Cartilage Intermediate Layer Protein 1 Suppresses TGF-β Signaling in Cardiac Fibroblasts

Background: Since transforming growth factor (TGF)-β1-induced cardiac fibrosis following myocardial infarction (MI) leads to heart failure and poor clinical prognosis, we aimed to identify a novel and unknown target for cardiac fibrosis related to the TGF-β signaling. Method and result: We performed and investigated RNA-Seq using infarcted mouse hearts, culminating in cartilage intermediate layer protein 1 (CILP1). Interestingly, Cilp1 expression was increased along with TGF-β1 expression in infarcted hearts, and was also upregulated after TGF-β1 stimulation in cardiac fibroblasts in vitro. Histological analysis revealed that Cilp1 was localized at the fibrotic regions of infarcted hearts. Full length CILP1 (F-CILP1) was cleaved into both N-terminal CILP1 (N-CILP1) and C-terminal CILP1 at the furin cleavage site, and both F-CILP1 and N-CILP1 were extracellularly secreted. We further found that CILP1 bound to TGF-β1 via thrombospondin type 1 domain, and suppressed both smad3 phosphorylation and fibroblasts differentiation to myofibroblasts induced by TGF-β1. Conclusion: We identified CILP1 as a potential regulator of cardiac fibrosis by inhibiting TGF-β signaling, and these results suggest the promise of CILP1 as a novel therapeutic target for preventing cardiac fibrosis and heart failure in MI patients

Increased endoplasmic reticulum stress in atherosclerotic plaques associated with acute coronary syndrome

BACKGROUND: The endoplasmic reticulum (ER) responds to various stresses by upregulation of ER chaperones, but prolonged ER stress eventually causes apoptosis. Although apoptosis is considered to be essential for the progression and rupture of atherosclerotic plaques, the influence of ER stress and apoptosis on rupture of unstable coronary plaques remains unclear. METHODS AND RESULTS: Coronary artery segments were obtained at autopsy from 71 patients, and atherectomy specimens were obtained from 40 patients. Smooth muscle cells and macrophages in the fibrous caps of thin-cap atheroma and ruptured plaques, but not in the fibrous caps of thick-cap atheroma and fibrous plaques, showed a marked increase of ER chaperone expression and apoptotic cells. ER chaperones also showed higher expression in atherectomy specimens from patients with unstable angina pectoris than in specimens from those with stable angina. Expression of 7-ketocholesterol was increased in the fibrous caps of thin-cap atheroma compared with thick-cap atheroma. Treatment of cultured coronary artery smooth muscle cells or THP-1 cells with 7-ketocholesterol induced upregulation of ER chaperones and apoptosis, whereas these changes were prevented by antioxidants. We also investigated possible signaling pathways for ER-initiated apoptosis and found that the CHOP (a transcription factor induced by ER stress)-dependent pathway was activated in unstable plaques. In addition, knockdown of CHOP expression by small interfering RNA decreased ER stress-dependent death of cultured coronary artery smooth muscle cells and THP-1 cells. CONCLUSIONS: Increased ER stress occurs in unstable plaques. Our findings suggest that ER stress-induced apoptosis of smooth muscle cells and macrophages may contribute to plaque vulnerability

Apoptosis inhibitor of macrophage depletion decreased M1 macrophage accumulation and the incidence of cardiac rupture after myocardial infarction in mice

<div><p>Background</p><p>Cardiac rupture is an important cause of death in the acute phase after myocardial infarction (MI). Macrophages play a pivotal role in cardiac remodeling after MI. Apoptosis inhibitor of macrophage (AIM) is secreted specifically by macrophages and contributes to macrophage accumulation in inflamed tissue by maintaining survival and recruiting macrophages. In this study, we evaluated the role of AIM in macrophage accumulation in the infarcted myocardium and cardiac rupture after MI.</p><p>Methods and results</p><p>Wild-type (WT) and AIM^‒/‒ mice underwent permanent left coronary artery ligation and were followed-up for 7 days. Macrophage accumulation and phenotypes (M1 pro-inflammatory macrophage or M2 anti-inflammatory macrophage) were evaluated by immunohistological analysis and RT-PCR. Matrix metalloproteinase (MMP) activity levels were measured by gelatin zymography. The survival rate was significantly higher (81.1% vs. 48.2%, <i>P</i><0.05), and the cardiac rupture rate was significantly lower in AIM^{<b>‒/‒</b>} mice than in WT mice (10.8% vs. 31.5%, <i>P</i><0.05). The number of M1 macrophages and the expression levels of M1 markers (iNOS and IL-6) in the infarcted myocardium were significantly lower in AIM^{<b>‒/‒</b>} mice than in WT mice. In contrast, there was no difference in the number of M2 macrophages and the expression of M2 markers (Arg-1, CD206 and TGF-β1) between the two groups. The ratio of apoptotic macrophages in the total macrophages was significantly higher in AIM^{<b>‒/‒</b>} mice than in WT mice, although MCP-1 expression did not differ between the two groups. MMP-2 and 9 activity levels in the infarcted myocardium were significantly lower in AIM^{<b>‒/‒</b>} mice than in WT mice.</p><p>Conclusions</p><p>These findings suggest that AIM depletion decreases the levels of M1 macrophages, which are a potent source of MMP-2 and 9, in the infarcted myocardium in the acute phase after MI by promoting macrophage apoptosis, and leads to a decrease in the incidence of cardiac rupture and improvements in survival rates.</p></div