Biology subjects

Ogawa, J.

Publications and source records attributed to Ogawa, J..

5 recordsLinked to original sources

bioRxiv2023

Synchronized long-read genome, methylome, epigenome, and transcriptome for resolving a Mendelian condition

Resolving the molecular basis of a Mendelian condition (MC) remains challenging owing to the diverse mechanisms by which genetic variants cause disease. To address this, we developed a synchronized long-read genome, methylome, epigenome, and transcriptome sequencing approach, which enables accurate single-nucleotide, insertion-deletion, and structural variant calling and diploid de novo genome assembly, and permits the simultaneous elucidation of haplotype-resolved CpG methylation, chromatin accessibility, and full-length transcript information in a single long-read sequencing run. Application of this approach to an Undiagnosed Diseases Network (UDN) participant with a chromosome X;13 balanced translocation of uncertain significance revealed that this translocation disrupted the functioning of four separate genes (NBEA, PDK3, MAB21L1, and RB1) previously associated with single-gene MCs. Notably, the function of each gene was disrupted via a distinct mechanism that required integration of the four omes to resolve. These included nonsense-mediated decay, fusion transcript formation, enhancer adoption, transcriptional readthrough silencing, and inappropriate X chromosome inactivation of autosomal genes. Overall, this highlights the utility of synchronized long-read multi-omic profiling for mechanistically resolving complex phenotypes.

genetics↗

bioRxiv2023

Discovery of an isoflavone oxidative catabolic pathway in legume root microbiota

Isoflavones are major specialized metabolites found in legume plants, where they contribute to environmental adaptation. Isoflavones also play a role human health as promising therapeutic agents. This metabolite group is involved in interactions with soil microorganisms as initiation signals in rhizobial symbiosis and as modulators of the legume root microbiota. We previously reported that isoflavones enrich the Comamonadaceae, a predominant bacterial family in soybean roots, and that microorganisms in legume rhizosphere soil degrade isoflavones. However, the isoflavone catabolism pathway that underly the isoflavone-mediated legume-microbiota interactions have not yet been clarified. Here, we isolated Variovorax sp. strain V35, member of the Comamonadaceae that harbors isoflavone-degrading activity, from soybean roots and discovered a gene cluster responsible for isoflavone degradation named ifc. Strain V35 metabolizes isoflavones in a completely distinct oxidative manner from the reductive isoflavone metabolism pathway elucidated in the gut microbiota, in which resulting products enter the tricarboxylic acid cycle. The characterization of ifc mutants and heterologously expressed IFC enzymes revealed that isoflavones are catabolized via A-ring cleaving fission, which starts with hydroxylation at the 8-position of the A-ring. We further demonstrated that ifc genes are frequently found in bacterial strains isolated from legume plants, including mutualistic rhizobia, and contribute to detoxification of the antibacterial activity of isoflavones. Taken together, our findings reveal an oxidative catabolism pathway of isoflavone in the soybean root microbiota, providing molecular insights into isoflavone-mediated legume-microbiota interactions. SignificanceIsoflavones play pivotal roles in plant-environment interactions and in the maintenance and improvement of human health. Bacterial metabolism is a fundamental component of isoflavone-mediated interkingdom interactions. In the human gut, intestinal bacteria convert isoflavones into equol, a highly bioactive compound. However, the fate of isoflavones in the legume rhizosphere has not been elucidated, despite them being the key signaling molecules for nodule symbiosis and modulation of the legume root microbiota. Here, we discovered a novel isoflavone catabolism pathway in the soybean root microbiota and demonstrated the strong association between bacterial catabolic abilities and their interactions with host plants. Collectively, our findings provide new insights into bacterial isoflavone metabolism and a molecular understanding of legume-microbiota interactions.

plant biology↗

bioRxiv2023

The gut lactic acid bacteria metabolite, 10-oxo-cis-6,trans-11-octadecadienoic acid, suppresses inflammatory bowel disease in mice by modulating the NRF2 pathway and GPCR-signaling

Various gut bacteria, including Lactobacillus plantarum, possess several enzymes that produce hydroxy fatty acids (FAs), oxo FAs, conjugated FAs, and partially saturated FAs from polyunsaturated FAs as secondary metabolites. Among these derivatives, we identified 10-oxo-cis-6,trans-11-octadecadienoic acid ({gamma}KetoC), a {gamma}-linolenic acid (GLA)-derived enon FA, as the most effective immunomodulator, which inhibited the antigen-induced immunoactivation and LPS-induced production of inflammatory cytokines. The treatment with {gamma}KetoC significantly suppressed proliferation of CD4+ T cells, LPS-induced activation of bone marrow-derived dendritic cells (BMDCs), and LPS-induced IL-6 release from peritoneal cells, splenocytes, and CD11c+ cells isolated from the spleen. {gamma}KetoC also inhibited the release of inflammatory cytokines from BMDCs stimulated with poly-I:C, R-848, or CpG. Further in vitro experiments using an agonist of GPR40/120 suggested the involvement of these GPCRs in the effects of {gamma}KetoC on DCs. We also found that {gamma}KetoC stimulated the NRF2 pathway in DCs, and the suppressive effects of {gamma}KetoC and agonist of GPR40/120 on the release of IL-6 and IL-12 were reduced in Nrf2-/- BMDCs. We evaluated the role of NRF2 in the anti-inflammatory effects of {gamma}KetoC in a dextran sodium sulfate-induced colitis model. The oral administration of {gamma}KetoC significantly reduced body weight loss, improved stool scores, and attenuated atrophy of the colon, in wild-type C57BL/6 and Nrf2+/- mice with colitis. In contrast, the pathology of colitis was deteriorated in Nrf2-/- mice even with the administration of {gamma}KetoC. Collectively, the present results demonstrated the involvement of the NRF2 pathway and GPCRs in {gamma}KetoC-mediated anti-inflammatory responses.

immunology↗

bioRxiv2022

Characterisation of an Escherichia coli line that completely lacks ribonucleotide reduction yields insights into the evolution of obligate intracellularity.

All life requires ribonucleotide reduction for de novo synthesis of deoxyribonucleotides. A handful of obligate intracellular species are known to lack ribonucleotide reduction and are instead dependent on their host for deoxyribonucleotide synthesis. As ribonucleotide reduction has on occasion been lost in obligate intracellular parasites and endosymbionts, we reasoned that it should in principle be possible to knock this process out entirely under conditions where deoxyribonucleotides are present in the growth media. We report here the creation of a strain of E. coli where all three ribonucleotide reductase operons have been fully deleted. Our strain is able to grow in the presence of deoxyribonucleosides and shows slowed but substantial growth. Under limiting deoxyribonucleoside levels, we observe a distinctive filamentous cell morphology, where cells grow but do not appear to divide regularly. Finally, we examined whether our lines are able to adapt to limited supplies of deoxyribonucleosides, as might occur in the evolutionary switch from de novo synthesis to dependence on host production during the evolution of parasitism or endosymbiosis. Over the course of an evolution experiment, we observe a 25-fold reduction in the minimum concentration of exogenous deoxyribonucleosides necessary for growth. Genome analysis of replicate lines reveals that several lines carry mutations in deoB and cdd. deoB codes for phosphopentomutase, a key part of the deoxyriboaldolase pathway, which has been hypothesised as an alternative to ribonucleotide reduction for deoxyribonucleotide synthesis. Rather than synthesis via this pathway complementing the loss of ribonucleotide reduction, our experiments reveal that mutations appear that reduce or eliminate the capacity for this pathway to catabolise deoxyribonucleotides, thus preventing their loss via central metabolism. Mutational inactivation of both deoB and cdd is also observed in a number of obligate intracellular bacteria that have lost ribonucleotide reduction. We conclude that our experiments recapitulate key evolutionary steps in the adaptation to intracellular life without ribonucleotide reduction.

microbiology↗

bioRxiv2020

The D614G mutation in the SARS-CoV2 Spike protein increases infectivity in an ACE2 receptor dependent manner

The SARS-CoV2 coronavirus responsible for the current COVID19 pandemic has been reported to have a relatively low mutation rate. Nevertheless, a few prevalent variants have arisen that give the appearance of undergoing positive selection as they are becoming increasingly widespread over time. Most prominent among these is the D614G amino acid substitution in the SARS-CoV2 Spike protein, which mediates viral entry. The D614G substitution, however, is in linkage disequilibrium with the ORF1b P314L mutation where both mutations almost invariably co-occur, making functional inferences problematic. In addition, the possibility of repeated new introductions of the mutant strain does not allow one to distinguish between a founder effect and an intrinsic genetic property of the virus. Here, we synthesized and expressed the WT and D614G variant SARS-Cov2 Spike protein, and report that using a SARS-CoV2 Spike protein pseudotyped lentiviral vector we observe that the D614G variant Spike has >1/2 log10 increased infectivity in human cells expressing the human ACE2 protein as the viral receptor. The increased binding/fusion activity of the D614G Spike protein was corroborated in a cell fusion assay using Spike and ACE2 proteins expressed in different cells. These results are consistent with the possibility that the Spike D614G mutant increases the infectivity of SARS-CoV2.

microbiology↗