Biology subjects

Nakajima, A.

Publications and source records attributed to Nakajima, A..

7 recordsLinked to original sources

bioRxiv2025

Uptake of fucosylated type I human milk oligosaccharide blocks by Bifidobacterium longum subsp. infantis

Human milk oligosaccharides (HMOs) are uniquely rich in the type 1 building block disaccharide lacto-N-biose I (LNB, Gal{beta}1,3GlcNAc), as compared to other mammals. Most HMOs are fucosylated, e.g., 1,2 and 1,4 fucosylations on LNB blocks, resulting in H type 1 (H1) and Lewis a (Lea) epitopes, respectively. The dominance of Bifidobacterium in breastfed infant guts hinges on efficient uptake of HMOs by specific ATP-binding cassette (ABC) importers. However, molecular insight into uptake of fucosylated LNB blocks is lacking. Here, we analyzed the uptake of LNB and its fucosylated H1 and Lea trisaccharides, as well as the mucin-derived disaccharide galacto-N-biose (GNB, Gal{beta}1,3GalNAc) by an ABC importer form the HMO-utilization specialist Bifidobacterium longum subsp. infantis. Structural analyses and molecular dynamics simulations explained how fucosylated and non-fucosylated LNB forms are recognized with similar affinities by the binding protein of this importer. Strikingly, we showed that two ABC importers confer to the uptake of LNB, while the Lea trisaccharide is efficiently internalized by a single importer in B. infantis. Phylogenetic and structural analyses of bifidobacterial ABC-associated binding proteins showed that the Lea clade harbors homologues possessing internal cavities, which allows for the accommodation of branched oligosaccharides. Our work provides unique insight into the evolution and molecular basis of capture and uptake of key HMO and host-derived saccharide blocks, highlighting these compounds as hitherto unexplored candidates for fortification of infant formula. ImportanceThe assembly of the gut microbiota in early life is critical to the health trajectory of human hosts. Breast feeding selects for a Bifidobacterium-rich community, adapted to efficiently utilize HMOs from mothers milk. Industrial scale production of HMOs for infant formula fortification has mainly considered fucosyllactoses, whereas fucosylated type 1 HMO blocks have hitherto not been explored. Our work sheds light on the uptake facet, central to the utilization of fucosylated HMOs with type 1 LNB building blocks. These type I blocks are efficiently internalized and assimilated by B. infantis, which has been recently shown to secrete immune-modulatory aromatic-lactate metabolites that mediate immune-priming of hosts in early life. This study contributes to our understanding of the utilization of HMOs and highlights fucosylated LNB blocks, as hitherto unexplored prebiotic candidates that support the growth of B. infantis and other beneficial bacteria in early life.

microbiology↗

bioRxiv2024

Traveling wave chemotaxis of neutrophil-like HL-60 cells

The question of how changes in chemoattractant concentration translate into the chemotactic response of immune cells serves as a paradigm for the quantitative understanding of how cells perceive and process temporal and spatial information. Here, using a microfluidic approach, we analyzed the migration of neutrophil-like HL-60 cells to a traveling wave of the chemoattractants fMLP and leukotriene B4 (LTB4). We found that under a pulsatile wave that travels at a speed of 95 and 170 {micro}m/min, cells move forward in the front of the wave but slow down and randomly orient at the back due to temporal decrease in the attractant concentration. Under a slower wave, cells re-orient and migrate at the back of the wave; thus, cell displacement is canceled out or even becomes negative as cells chase the receding wave. FRET-based analysis indicated that these patterns of movement correlated well with spatiotemporal changes in Cdc42 activity. Furthermore, pharmacological perturbations suggested that migration in front of the wave depends on Cdc42, whereas that in the back of the wave depends more on PI3K/Rac and ROCK. These results suggest that pulsatile attractant waves may recruit or disperse neutrophils, depending on their speed and degree of cell polarization. Significance statementO_LIThe way neutrophil chemotaxis is directed by attractants is thought to depend on temporal and spatial concentration changes, however the response to a transient pulsatile stimulus has not been well explored. C_LIO_LIA moderately fast traveling wave of fMLP and leukotrieneB4 (LTB4) directs unidirectional cell migration. Under slow waves, cells respond to the back of the wave; thus, cell displacement is canceled out. Cdc42, PI3K, Rac, and ROCK contributed differently to responses at the front and back of the wave. C_LIO_LIThese findings suggest that traveling waves of attractant can guide immune cell recruitment and dispersal. C_LI

cell biology↗

bioRxiv2022

Human Milk Oligosaccharide Utilization in Intestinal Bifidobacteria is Governed by a Global Transcriptional Regulator NagR

Bifidobacterium longum subsp. infantis (B. infantis) is a prevalent beneficial bacterium that colonizes the human neonatal gut and is uniquely adapted to efficiently use human milk oligosaccharides (HMOs) as a carbon and energy source. Multiple studies have focused on characterizing the elements of HMO utilization machinery in B. infantis; however, the regulatory mechanisms governing the expression of these catabolic pathways remain poorly understood. A bioinformatic regulon reconstruction approach used in this study implicated NagR, a transcription factor from the ROK family, as a negative global regulator of genomic loci encoding lacto-N-biose/galacto-N-biose (LNB/GNB), lacto-N-tetraose (LNT), and lacto-N-neotetraose (LNnT) utilization pathways in B. infantis. This conjecture was corroborated by transcriptome profiling upon nagR genetic inactivation and experimental assessment of binding of recombinant NagR to predicted DNA operators. The latter approach also implicated N-acetylglucosamine (GlcNAc), a universal intermediate of LNT and LNnT catabolism, and its phosphorylated derivatives as plausible NagR transcriptional effectors. Reconstruction of NagR regulons in various Bifidobacterium lineages revealed multiple regulon expansion events, suggesting evolution from a local regulator of GlcNAc catabolism in ancestral bifidobacteria to a global regulator controlling foraging of mixtures of GlcNAc-containing host-derived glycans in mammalian gut-colonizing B. infantis and Bifidobacterium bifidum. ImportanceThe predominance of bifidobacteria in the gut of breastfed infants is attributed to the ability of these bacteria to utilize human milk oligosaccharides (HMOs). Thus, individual HMOs such as lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT) are considered promising prebiotics that would stimulate the growth of bifidobacteria and confer multiple health benefits to preterm and malnourished children suffering from impaired (stunted) gut microbiota development. However, the rational selection of HMO-based prebiotics is hampered by the incomplete knowledge of regulatory mechanisms governing HMO utilization in target bifidobacteria. This study describes NagR-mediated transcriptional regulation of LNT and LNnT utilization in Bifidobacterium longum subsp. infantis. The elucidated regulatory network appears optimally adapted to simultaneous utilization of multiple HMOs, providing a rationale to add HMO mixtures (rather than individual components) into infant formulas. The study also provides insights into the evolutionary trajectories of complex regulatory networks controlling carbohydrate metabolism in bifidobacteria.

microbiology↗

bioRxiv2021

The impact of PNPLA3 and TM6SF2 in cirrhosis related complications

Patatin-like phospholipase domain-containing 3 (PNPLA3) and transmembrane 6-superfamily member 2 (TM6SF2) polymorphisms have major impact for fibrosis due to steatohepatitis. However, there are scant data about correlations between cirrhosis-related complications and the polymorphisms of these genes. Therefore, we aimed to determine the role of the PNPLA3 and TM6SF2 polymorphisms in fibrosis progression for patients with liver cirrhosis. A multicenter study was performed at six hospitals in Japan enrolling 400 patients with liver cirrhosis caused by virus (n = 157), alcohol (n = 104), nonalcoholic fatty liver disease (NAFLD) (n = 106), or autoimmune disease (n = 33). These cirrhotic patients included those with complications of variceal bleeding, hepatic ascites, and/or hepatic encephalopathy and those without. To assess the role of the PNPLA3 and TM6SF2 polymorphisms in patients with cirrhosis related complications, we calculated the odds ratio and relative risk for the rs738409 and rs58542926 polymorphisms. We also accessed whether or not the interaction between these two polymorphisms contributed to cirrhosis related complications. As a result, the odds ratio for complications in the NAFLD group significantly increased in the presence of the rs738409 GG genotype when the CC genotype was used as the reference. There were no significant risks between complications and the presence of the rs738409 G allele in the virus or alcohol groups. There were no significant risks of complications in the frequency of the rs58542926 T polymorphism regardless of the etiology of liver cirrhosis. The interaction between the rs738409 and rs58542926 polymorphisms had the highest odds ratio of 2.415 for complications in the rs738409 GG + rs58542926 (CT+TT) group when rs738409 (CC+CG) + TM6SF2 CC was used as the reference in the NAFLD group although there was no statistically interaction between these gene polymorphisms (P=0.085).

genetics↗

bioRxiv2020

Comparative mapping of crawling-cell morphodynamics in deep learning-based feature space

Navigation of fast migrating cells such as amoeba Dictyostelium and immune cells are tightly associated with their morphologies that range from steady polarized forms that support high directionality to those more complex and variable when making frequent turns. Model simulations are essential for quantitative understanding of these features and their origins, however systematic comparisons with real data are underdeveloped. Here, by employing deep-learning-based feature extraction combined with phase-field modeling framework, we show that a low dimensional feature space for 2D migrating cell morphologies obtained from the shape stereotype of keratocytes, Dictyostelium and neutrophils can be fully mapped by interlinked signaling network of cell-polarization and protrusion dynamics. Our analysis links the data-driven shape analysis to the underlying causalities by identifying key parameters critical for migratory morphologies both normal and aberrant under genetic and pharmacological perturbations. The results underscore the importance of deciphering self-organizing states and their interplay when characterizing morphological phenotypes.

cell biology↗

bioRxiv2020

Micro-topographical guidance of macropinocytic signaling patches

In fast moving cells such as amoeba and immune cells, spatio-temporal regulation of dendritic actin filaments shapes large-scale plasma membrane protrusions. Despite the importance in migration as well as in particle and liquid ingestion, how these processes are affected by the micrometer-scale surface features is poorly understood. Here, through quantitative imaging analysis of Dictyostelium on micro-fabricated surfaces, we show that there is a distinct mode of topographically guided cell migration phagotaxis directed by the macropinocytic Ras/PI3K signaling patches. The topography guidance was PI3K-dependent and involved nucleation of a patch at the convex curved surface and confinement at the concave surface. Due to the topography-dependence, constitutive cup formation for liquid uptake in the axenic strain is also destined to trace large surface features. Given the fact that PI3K-dependency of phagocytosis are restricted to large particles in both Dictyostelium and immune cells, topography-dependency and the dual-use of membrane cups may be wide-spread.

biophysics↗

bioRxiv2018

Tissue self-organization based on collective cell migration by contact activation of locomotion and chemotaxis

Despite their central role in multicellular organization, navigation rules that dictate cell rearrangement remain much to be elucidated. Contact between neighboring cells and diffusive attractant molecules are two of the major determinants of tissue-level patterning, however in most cases, molecular and developmental complexity hinders one from decoding the exact governing rules of individual cell movement. A primordial example of tissue patterning by cell rearrangement is found in the social amoeba Dictyostelium discoideum where the organizing center or the tip self-organize as a result of sorting of differentiating prestalk and prespore cells. Due to its relatively simple and conditional multicellularity, the system provides a rare case where the process can be fully dissected into individual cell behavior. By employing microfluidics and microsphere-based manipulation of navigational cues at the single-cell level, here we uncovered a previously overlooked mode of Dictyostelium cell migration that is strictly directed by cell-cell contact. The cell-cell contact signal is mediated by E-set Ig-like domain containing heterophilic adhesion molecules TgrB1/TgrC1 that act in trans to induce plasma membrane recruitment of SCAR complex and formation of dendritic actin networks, and the resulting cell protrusion competes with those induced by chemoattractant cAMP. Furthermore, we demonstrate that both prestalk and prespore cells can protrude towards the contact signal as well as to chemotax towards cAMP, however when given both signals, prestalk cells orient towards the chemoattractant whereas prespore cells choose the contact signal. These data suggest a new model of cell sorting by competing juxtacrine and diffusive cues each with potential to drive its own mode of collective cell migration. The present findings not only resolve the long standing question of how cells sort in Dictyostelium but also cast light on the remarkable parallels in collective cell migration that evolved independently in metazoa and amoebozoa.

cell biology↗