Biology subjects

Nagata, Y.

Publications and source records attributed to Nagata, Y..

5 recordsLinked to original sources

bioRxiv2026

Entry exclusion enables selective conjugative DNA delivery in synthetic bacterial communities.

Selective DNA delivery to specific members of assembled bacterial communities remains challenging. Bacterial conjugation enables efficient DNA delivery, but transfer to non-target recipients limits its specificity within mixed communities. Here, we repurpose plasmid entry exclusion (Eex) as a recipient-side gate to control conjugative DNA delivery. We demonstrate selective plasmid delivery to Eex-negative recipients within populations containing both Eex-expressing and Eex-negative cells. This recipient selectivity was maintained at increased cell densities and during prolonged mating. By combining RP4-type and F-type conjugation systems with their corresponding exclusion modules, we further directed DNA delivery from distinct donors to defined recipient populations. RP4-derived Eex also functioned in environmental bacteria, including Pseudomonas putida and Sphingobium japonicum, enabling recipient-specific exclusion within a multispecies mixture. Furthermore, repeated cycles of Eex-guided conjugation and selection altered community composition after assembly. These results establish entry exclusion as a recipient-side strategy for selective conjugative DNA delivery and, when combined with selection, for controlling the composition of assembled bacterial communities.

microbiology↗

bioRxiv2026

Average local nucleosome motion remains constant during interphase in living human cells

BackgroundDynamic chromatin behavior, which is related to chromatin accessibility, plays a critical role in various genome DNA functions such as RNA transcription and DNA replication/repair. Previous studies using highly synchronized cells showed that average local chromatin motion, captured by single-nucleosome imaging and tracking on a second time scale, remained almost constant throughout G1, S, and G2 phases in living human cells, although possible effects of prolonged drug treatments for cell-cycle synchronization could not be excluded. ResultsTo avoid possible effects of prolonged drug treatment, we combined single-nucleosome imaging with Fucci probes to visualize cell-cycle progression through G1, S, and G2. Using HeLa and HCT116 cells expressing H2B-HaloTag and Fucci probes, we found that local nucleosome motion remained similar on average throughout interphase, except for elevated motion in early G1. Transcription inhibition similarly increased nucleosome motion throughout interphase. Local nucleosome motion also increased following replication stress or DNA damage. ConclusionOur findings suggest that near-constant chromatin motion supports housekeeping functions under similar physical conditions during interphase. Our findings also suggest that cells can transiently change chromatin motion to perform ad hoc tasks in response to signals from inside and outside the cell, such as DNA damage.

cell biology↗

bioRxiv2025

Type I partition-related proteins enhance conjugative transfer through transcriptional activation and oriT region binding

Plasmid partitioning and bacterial conjugation are critical processes ensuring plasmid maintenance and dissemination, respectively, within bacterial populations. Although traditionally regarded as distinct phenomena, these two processes are increasingly recognized as interconnected. While partitioning ensures plasmid inheritance during cell division, its potential influence on conjugative transfer remains poorly understood. A major impediment to understanding their interplay is that partition systems are often essential for plasmid stability, making it difficult to distinguish their direct effects on conjugation. In this study, we addressed this challenge using a mini-conjugative plasmid derived from the Pseudomonas putida NAH7 plasmid. This engineered plasmid, containing all conjugation-related genes, was cloned into an E. coli-compatible vector. Additionally, the par genes from NAH7 were expressed from a separate plasmid to investigate their roles in conjugative transfer. Our results revealed that the par gene cluster plays a significant role in enhancing the conjugative transfer of the mini-conjugative plasmid. Specifically, ParB, a centromere-binding protein, functions as a transcriptional activator of conjugation-related genes with binding parSNAH site. In contrast, ParR, a KorA homolog, was not found to enhance transcription directly but binds extensively to the oriT region. This binding probably facilitates the recruitment or stabilization of the relaxosome, thereby enhancing conjugation efficiency. Together, these findings unveil a previously unappreciated role for partition proteins in stimulating bacterial conjugation, providing new insights into how plasmids coordinate vertical and horizontal dissemination, highlighting that these processes can occur simultaneously within bacterial communities. IMPORTANCEPlasmid partition systems are classified into three types. While some systems have been reported to influence conjugative transfer, this study uncovers a novel mechanism utilized by a Type I system to enhance DNA transfer. Strikingly, repeat sequences perfectly matching the parSNAH site--bound by ParB to activate downstream conjugative transfer genes--were identified on both plasmids and chromosomes across diverse proteobacterial taxa. Furthermore, many of these repeat sequences were localized near genes involved in conjugative transfer and partitioning, suggesting the presence of a conserved regulatory mechanism mediated by these repeats. This study provides important insights into how plasmid partition systems coordinate both vertical and horizontal dissemination. Such knowledge is essential for understanding and mitigating the spread of antibiotic resistance and other plasmid-encoded traits, and it offers a foundation for developing strategies to manage plasmid-associated genetic exchange.

microbiology↗

bioRxiv2024

Evolution of Tn4371 family ICE; traR mediated coordination of cargo gene upregulation and horizontal transfer

ICEKKS102Tn4677, which has been shown to transfer horizontally, carries bph operon for mineralization of PCBs/biphenyl and belongs to an ICE Tn4371 family. In this study we investigated the role of traR gene encoding a LysR-type transcriptional regulator, which is conserved in sequence, positioning, and directional orientation among Tn4371 family ICEs. The traR belonged to bph operon and its overexpression on solid medium resulted in modest upregulation of traG (3-fold) and marked upregulation of xis (80-fold), and enhanced ICE excision, and notably ICE transfer frequency. We propose the evolutional roles of traR, which upon insertion to the current position, connected the cargo gene activation and ICE-transfer. This property of ICE, transferring under environmental conditions that lead to cargo gene activation, would give fitness advantages to the host bacteria, thereby resulting in efficient dissemination of the Tn4371 family ICEs. SignificanceOnly ICEKKS102Tn4677 is proven to transfer among the widely disseminating Tn4371 family ICEs from {beta} and {gamma}-proteobacteria. We showed that the traR gene in ICEKKS102Tn4677 conserved in the ICE family with fixed location and direction is co-transcribed with the cargo gene and activates ICE transfer. We propose that capturing of traR by an ancestral ICE to the current position established ICE Tn4371 family ICEs. Our findings provide insights into the evolutionary processes that led to the widespread distribution of the Tn4371 family of ICEs across bacterial species.

molecular biology↗

bioRxiv2019

Nintedanib ameliorates experimental pulmonary arterial hypertension via inhibition of endothelial mesenchymal transition and smooth muscle cell proliferation

Neointimal lesion and medial wall thickness of pulmonary arteries (PAs) are common pathological findings in pulmonary arterial hypertension (PAH). Platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF) signaling contribute to intimal and medial vascular remodeling in PAH. Nintedanib is a tyrosine kinase inhibitor whose targets include PDGF and FGF receptors. Although the beneficial effects of nintedanib were demonstrated for human idiopathic pulmonary fibrosis, its efficacy for PAH is still unclear. Thus, we hypothesized that nintedanib is a novel treatment for PAH to inhibit the progression of vascular remodeling in PAs. The inhibitory effects of nintedanib were evaluated both in endothelial mesenchymal transition (EndMT)-induced human pulmonary microvascular endothelial cells (HPMVECs) and human pulmonary arterial smooth muscle cells (HPASMCs) stimulated by growth factors. We also tested the effect of chronic nintedanib administration on a PAH rat model induced by Sugen5416 (a VEGF receptor inhibitor) combined with chronic hypoxia. Nintedanib was administered from weeks 3 to 5 after Sugen5416 injection, and pulmonary hemodynamics and PAs pathology were evaluated. Nintedanib attenuated the expression of mesenchymal markers in EndMT-induced HPMVECs and HPASMCs proliferation. Phosphorylation of PDGF and FGF receptors was augmented both in both intimal and medial lesions of PAs. Nintedanib blocked these phosphorylation, improved hemodynamics and reduced vascular remodeling involving neointimal lesions and medial wall thickening in PAs. Additionally, expressions Twist1, transcription factors associated with EndMT, in lung tissue was significantly reduced by nintedanib. These results suggest that nintedanib may be a novel treatment for PAH with anti-vascular remodeling effects.

physiology↗