Biology subjects

Naito, S.

Publications and source records attributed to Naito, S..

7 recordsLinked to original sources

bioRxiv2024

Nobiletin, a Polymethoxyflavonoid, Activates the Desuccinylase Activity of SIRT5 and Prevents the Development of Heart Failure

Nobiletin is a natural compound useful for the prevention and treatment of several diseases. However, the precise role of nobiletin in heart failure is unclear. Nobiletin treatment prevents pressure overload- and myocardial infarction-induced heart failure. Using affinity purification of biotinylated nobiletin from rat heart cell lysates, we identified sirtuin 5 (SIRT5) as a novel nobiletin-binding protein. Nobiletin enhanced the desuccinylase activity of SIRT5 in vitro. Compared to wild-type mice, SIRT5-overexpressing transgenic mice resisted pressure overload-induced systolic dysfunction. Conversely, SIRT5 knockout disrupted the nobiletin-mediated therapeutic effects on heart failure in mice. SIRT5 desuccinylated p300 at lysine 1568 and reduced the histone acetyltransferase (HAT) activity of p300. The desuccinylated p300 mutant suppressed the phenylephrine-induced cardiomyocyte hypertrophic responses. These findings suggest that nobiletin prevents heart failure development through SIRT5-dependent inhibition of p300-HAT activity. Nobiletin, a nontoxic dietary compound, is a potential therapeutic agent for heart failure in humans.

molecular biology↗

bioRxiv2022

The conserved upstream ORF of the Arabidopsis ANAC082 gene mediates translational upregulation in response to nucleolar stress

Perturbations in ribosome biogenesis cause a type of cellular stress called nucleolar or ribosomal stress, which triggers adaptive responses in both animal and plant cells. The Arabidopsis ANAC082 transcription factor has been identified as a key mediator of the plant nucleolar stress response. The 5'-untranslated region (5'-UTR) of ANAC082 mRNA contains an upstream ORF (uORF) encoding an evolutionarily conserved amino acid sequence. Here, we report that this uORF mediates the upregulation of ANAC082 translation in response to nucleolar stress. When transgenic Arabidopsis plants containing a luciferase reporter gene under the control of the ANAC082 promoter and 5'-UTR were treated with reagents that induced nucleolar stress, translation of the reporter gene was enhanced in a uORF sequence-dependent manner. Additionally, we examined the effect of an endoplasmic reticulum (ER) stress-inducing reagent on reporter gene expression because the closest homolog of ANAC082 in Arabidopsis, ANAC103, is involved in the ER stress response. However, the ANAC082 uORF did not respond to ER stress. Interestingly, although ANAC103 has a uORF with an amino acid sequence similar to that of the ANAC082 uORF, the C-terminal sequence critical for regulation is not well conserved among ANAC103 homologs in Brassicaceae. Transient expression assays revealed that unlike the ANAC082 uORF, the ANAC103 uORF does not exert a sequence-dependent regulatory effect. Altogether, our findings suggest that the ANAC082 uORF is important for the nucleolar stress response but not for the ER stress response, and that for this reason, the uORF sequence-dependent translational regulation was lost in ANAC103 during evolution.

plant biology↗

bioRxiv2021

Placental mammals acquired the functional region and domain in NRK for regulating the CK2-PTEN-AKT pathway and placental cell proliferation

The molecular evolution processes underlying the acquisition of the placenta in eutherian ancestors are not fully understood. Mouse NCK-interacting kinase (NIK)-related kinase (NRK) is expressed highly in the placenta and plays a role in preventing placental hyperplasia. Here, we show the molecular evolution of NRK, which confers its function for inhibiting placental cell proliferation. Comparative genome analysis identified NRK orthologues across vertebrates, which share the kinase and citron homology (CNH) domains. Evolutionary analysis revealed that NRK underwent extensive amino acid substitutions in the ancestor of placental mammals and has been since conserved. Biochemical analysis of mouse NRK revealed that the CNH domain binds to phospholipids, and a region in NRK binds to and inhibits casein kinase-2 (CK2), which we named the CK2-inhibitory region (CIR). Cell culture experiments suggest the following: (1) mouse NRK is localised at the plasma membrane via the CNH domain, where the CIR inhibits CK2. (2) This mitigates CK2-dependent phosphorylation and inhibition of PTEN, and (3) leads to the inhibition of AKT signalling and cell proliferation. Nrk deficiency increased phosphorylation levels of PTEN and AKT in mouse placenta, supporting our hypothesis. Unlike mouse NRK, chicken NRK did not bind to phospholipids and CK2, decrease phosphorylation of AKT, or inhibit cell proliferation. Both the CNH domain and CIR have evolved under purifying selection in placental mammals. Taken together, our study suggests that placental mammals acquired the phospholipid-binding CNH domain and CIR in NRK for regulating the CK2-PTEN-AKT pathway and placental cell proliferation.

molecular biology↗

bioRxiv2021

Genome-wide identification of Arabidopsis non-AUG-initiated upstream ORFs with evolutionarily conserved regulatory sequences that control protein expression levels

Upstream open reading frames (uORFs) are short ORFs found in the 5'-UTRs of many eukaryotic transcripts and can influence the translation of protein-coding main ORFs (mORFs). Recent genome-wide ribosome profiling studies have revealed that thousands of uORFs initiate translation at non-AUG start codons. However, the physiological significance of these non-AUG uORFs has so far been demonstrated for only a few of them. It is conceivable that physiologically important non-AUG uORFs are evolutionarily conserved across species. In this study, using a combination of bioinformatics and experimental approaches, we searched the Arabidopsis genome for non-AUG-initiated uORFs with conserved sequences that control the expression of the mORF-encoded proteins. As a result, we identified four novel regulatory non-AUG uORFs. Among these, two exerted repressive effects on mORF expression in an amino acid sequence-dependent manner. These two non-AUG uORFs are likely to encode regulatory peptides that cause ribosome stalling, thereby enhancing their repressive effects. In contrast, one of the identified regulatory non-AUG uORFs promoted mORF expression by alleviating the inhibitory effect of a downstream AUG-initiated uORF. These findings provide insights into the mechanisms that enable non-AUG uORFs to play regulatory roles despite their low translation initiation efficiencies.

genomics↗

bioRxiv2019

Exhaustive identification of conserved upstream open reading frames with potential translational regulatory functions from animal genomes

Upstream open reading frames (uORFs) are present in the 5-untranslated regions of many eukaryotic mRNAs, and some peptides encoded by these regions play important regulatory roles in controlling main ORF (mORF) translation. We previously developed a novel pipeline, ESUCA, to comprehensively identify plant uORFs encoding functional peptides, based on genome-wide identification of uORFs with conserved peptide sequences (CPuORFs). Here, we applied ESUCA to diverse animal genomes, because animal CPuORFs have been identified only by comparing uORF sequences between a limited number of species, and how many previously identified CPuORFs encode regulatory peptides is unclear. By using ESUCA, 1,517 (1,373 novel and 144 known) CPuORFs were extracted from four evolutionarily divergent animal genomes. We examined the effects of 17 human CPuORFs on mORF translation using transient expression assays. Through these analyses, we identified seven novel regulatory CPuORFs that repressed mORF translation in a sequence-dependent manner, including one conserved only among Eutheria. We discovered a much higher number of animal CPuORFs than previously identified. Since most human CPuORFs identified in this study are conserved across a wide range of Eutheria or a wider taxonomic range, many CPuORFs encoding regulatory peptides are expected to be found in the identified CPuORFs.

bioinformatics↗

bioRxiv2019

ESUCA: a pipeline for genome-wide identification of upstream open reading frames with evolutionarily conserved sequences and determination of the taxonomic range of their conservation

BackgroundUpstream open reading frames (uORFs) in the 5'-untranslated regions (5'-UTRs) of certain eukaryotic mRNAs encode evolutionarily conserved functional peptides, such as cis-acting regulatory peptides that control translation of downstream main ORFs (mORFs). For genome-wide searches for uORFs with conserved peptide sequences (CPuORFs), comparative genomic studies have been conducted, in which uORF sequences were compared between selected species. To increase chances of identifying CPuORFs, we previously developed an approach in which uORF sequences were compared using BLAST between Arabidopsis and any other plant species with available transcript sequence databases. If this approach is applied to multiple plant species belonging to phylogenetically distant clades, it is expected to further comprehensively identify CPuORFs conserved in various plant lineages, including those conserved among relatively small taxonomic groups.\n\nResultsTo efficiently compare uORF sequences among many species and efficiently identify CPuORFs conserved in various taxonomic lineages, we developed a novel pipeline, ESUCA. We applied ESUCA to the genomes of five angiosperm species, which belong to phylogenetically distant clades, and selected CPuORFs conserved among at least three different orders. Through these analyses, we identified 88 novel CPuORF families. As expected, ESUCA analysis of each of the five angiosperm genomes identified many CPuORFs that were not identified from ESUCA analyses of the other four species. However, unexpectedly, these CPuORFs include those conserved in wide taxonomic ranges, indicating that the approach used here is useful not only for comprehensive identification of narrowly conserved CPuORFs but also for that of widely conserved CPuORFs. Examination of the effects of 11 selected CPuORFs on mORF translation revealed that CPuORFs conserved only in relatively narrow taxonomic ranges can have sequence-dependent regulatory effects, suggesting that most of the identified CPuORFs are conserved because of functional constraints of their encoded peptides.\n\nConclusionsThis study demonstrates that ESUCA is capable of efficiently identifying CPuORFs likely to be conserved because of the functional importance of their encoded peptides. Furthermore, our data show that the approach in which uORF sequences from multiple species are compared with those of many other species, using ESUCA, is highly effective in comprehensively identifying CPuORFs conserved in various taxonomic ranges.

bioinformatics↗

bioRxiv2018

Galectin-1 promotes the invasion of bladder cancer urothelia through their matrix milieu

The progression of carcinoma of the urinary bladder involves migration of cancer epithelia through their surrounding tissue matrix microenvironment. This was experimentally confirmed when a gender- and grade-diverse set of bladder cancer cell lines were cultured in pathomimetic three-dimensional laminin-rich environments. The high-grade cells, particularly female, formed multicellular invasive morphologies in 3D. In comparison, low- and intermediate-grade counterparts showed growth-restricted phenotypes. A proteomic approach combining mass spectrometry and bioinformatics analysis identified the estrogen-driven lactose-binding lectin Galectin-1 (GAL-1) as a putative candidate that could drive this invasion. Expression of LGALS1, the gene encoding GAL-1 showed an association with tumor grade progression in bladder cell lines. Immunohisto- and cyto-chemical experiments suggested greater extracellular levels of GAL-1 in 3D cultures of high-grade bladder cells and cancer tissues. High levels of GAL-1 associated with increased proliferation- and adhesion- of bladder cancer cells when grown on laminin-rich matrices. Pharmacological inhibition and Gal-1 knockdown in high-grade female cells decreased their adhesion to, and viability on, laminin-rich substrata. Higher GAL-1 also correlated with reduced E-cadherin and increased N-cadherin levels in consonance with a mesenchymal-like phenotype that we observed in 3D culture. The inhibition of GAL-1 reversed the stellate invasive phenotype to a more growth-restricted one in high-grade cells embedded within both basement-membrane-like and stromal collagenous matrix scaffolds. Finally, inhibition of GAL-1 specifically altered cell surface sialic acids, suggesting the mechanism by which the levels of GAL-1 may underlie the aggression and poor prognosis of invasive bladder cancer, especially in women.

cancer biology↗