Biology subjects

Tobita, H.

Publications and source records attributed to Tobita, H..

5 recordsLinked to original sources

bioRxiv2025

Evolution of circadian clock and light-input pathway genes in Hemiptera

Circadian clocks are timekeeping mechanisms that help organisms anticipate periodic alterations of day and night. These clocks are widespread, and in the case of animals, they rely on genetically related components. At the molecular level, the animal circadian clock consists of several interconnected transcription-translation feedback loops. Although the clock setup is generally conserved, some important differences exist even among various insect groups. Therefore, we decided to identify in silico all major clock components and closely related genes in Hemiptera. Our analyses indicate several lineage-specific alterations of the clock setup in Hemiptera, derived from gene losses observed in the complete gene set identified in the outgroup, Thysanoptera, which thus presents the insect lineage with a complete clock setup. Nilaparvata and Fulgoroidea, in general, lost the (6-4) photolyase, while all Hemiptera lost FBXL3, and several lineage-specific losses of dCRY and jetlag were identified. Importantly, we identified non-canonical splicing variants of period and m-cry genes, which might provide another regulatory mechanism for clock functioning. Lastly, we performed a detailed reconstruction of Hemipteras light input pathway genetic repertoire and explored the horizontal gene transfer of cryptochrome-DASH from plant to Bemisia. Altogether, this inventory reveals important trends in clock gene evolution and provides a reference for clock research in Hemiptera, including several lineages of important pest species. Bullet points (highlights){circ} Evolution of clock genes (including light input pathway) was reconstructed in Hemiptera {circ}New m-cry and per splicing variants were identified in certain species {circ}A unique horizontal gene transfer of plant/fungal CRY-DASH was found in Bemisia {circ}Clock setup was identified for pests: Nilaparvata, Bemisia, Halyomorpha, and aphids {circ}Future clock research directions in Hemiptera are proposed

evolutionary biology↗

bioRxiv2024

Knockout of cryptochrome 1 disrupts circadian rhythm and photoperiodic diapause induction in the silkworm, Bombyx mori

Most insects enter diapause, a state of physiological dormancy crucial for enduring harsh seasons, with photoperiod serving as the primary cue for its induction, ensuring proper seasonal timing of the process. Although the involvement of the circadian clock in the photoperiodic time measurement has been demonstrated through knockdown or knockout of clock genes, the precise molecular mechanisms in this context remain unclear. In bivoltine strains of the silkworm, Bombyx mori, embryonic diapause is maternally controlled and affected by environmental conditions experienced by mother moths during embryonic and larval stages. Previous research highlighted the role of core clock genes, including period (per), timeless (tim), Clock (Clk) and cycle (cyc), in photoperiodic diapause induction in B. mori. In this study, we focused on another clock gene, cryptochrome 1 (cry1), which functions as a photoreceptor implicated in photoentrainment of the circadian clock across various insect species. Phylogenetic analysis and conserved domain identification confirmed the presence of both Drosophila-type cry (cry1) and mammalian-type cry (cry2) genes in the B. mori genome, akin to other lepidopterans. Temporal expression analysis revealed higher cry1 gene expression during the photophase and lower expression during the scotophase, with knockouts of core clock genes (per, tim, Clk and cyc) disrupting this temporal expression pattern. Using CRISPR/Cas9-mediated genome editing, we established a cry1 knockout strain in p50T, a bivoltine strain exhibiting clear photoperiodism during both embryonic and larval stages. Although the wild-type strain displayed circadian rhythm in eclosion under continuous darkness, the cry1 knockout strain exhibited arrhythmic eclosion, implicating B. mori cry1 in the circadian clock feedback loop governing behavior rhythms. Females of the cry1 knockout strain failed to induce photoperiodic diapause during both embryonic and larval stages, mirroring the diapause phenotype of the wild-type individuals reared under constant darkness, indicating that B. mori CRY1 contributes to photoperiodic time measurement as a photoreceptor. Furthermore, photoperiodic diapause induction during the larval stage was abolished in a cry1/tim double-knockout strain, suggesting that photic information received by CRY1 is relayed to the circadian clock. Overall, this study represents the first evidence of cry1 involvement in insect photoperiodism, specifically in diapause induction. HighlightsO_LIKnockouts of core clock genes disrupted the rhythmic expression of cryptochrome 1 (cry1). C_LIO_LIA cry1 knockout strain was established using CRISPR/Cas9. C_LIO_LIThe cry1 knockout strain lost its eclosion rhythm. C_LIO_LIKnockout of cry1 disrupted photoperiodic diapause induction. C_LIO_LIFemales of a cry1/tim double knockout strain produced only non-diapause eggs regardless of larval photoperiod. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/593801v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@1a2b363org.highwire.dtl.DTLVardef@af3adcorg.highwire.dtl.DTLVardef@41490corg.highwire.dtl.DTLVardef@22bc30_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

bioRxiv2022

Knockouts of positive and negative elements of the circadian clock disrupt photoperiodic diapause induction in the silkworm, Bombyx mori

Diapause is one of the most important traits that have sustained insects to thrive. Insects can prospectively arrest their development and increase environmental stress-resistance by entering diapause. The photoperiod is the signal that indicates insects the proper timing to enter diapause. Circadian clock genes are shown to be involved in photoperiodic diapause induction in various insect species. The silkworm, Bombyx mori, enters diapause at the embryonic stage. In bivoltine strains, diapause determination is affected by embryonic temperature and photoperiodic conditions during embryonic and larval stages. Two independent studies showed that knocking out the core clock gene, period, perturb photoperiodic diapause induction. However, whether the circadian clock as whole or individual clock genes are responsible for the photoperiodic diapause induction remains unknown. In this study, using CRISPR/Cas9 we knocked out negative (period and timeless) and positive elements (Clock and cycle) in p50T, a bivoltine strain which exhibits photoperiodic diapause induction during both embryonic and larval stages. The temporal expression patterns of clock genes changed in each core clock gene knockout strain, suggesting disruption of normal feedback loops produced by circadian clock genes. Furthermore, photoperiodic diapause induction during both embryonic and larval stages was lost in all knockout strains. Our results indicate the involvement of circadian clock in photoperiodic diapause induction in B. mori. HighlightsO_LIWe established knockout Bombyx mori strains of four core clock genes C_LIO_LIThe temporal expression patterns of clock genes changed in knockout strains C_LIO_LIPhotoperiodic diapause induction was not observed in any knockout strains C_LI Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

genetics↗

bioRxiv2022

Conifer leaves have a peroxisomal oxidative decarboxylation path in the photorespiratory pathway

The photorespiratory pathway consists of enzymes operating in chloroplasts, mitochondria, and peroxisomes. Conifer leaves lack one of them, chloroplastic Gln synthetase, which questioned the current belief that the photorespiratory mechanism is identical between angiosperm C3 species and conifers. A photorespiratory-metabolite analysis of the leaves of 13 conifer and 14 angiosperm tree species revealed significant differences in the mean metabolite concentrations between the two taxonomic groups: the glycerate content on chlorophyll basis in conifer leaves was <1/10 that detected in angiosperm leaves, whereas the glycolate content was 1.6 times higher in conifer leaves. Glycerate is produced from Ser through an intermediate, hydroxypyruvate. To investigate the lower glycerate levels observed in conifer leaves, we performed experiments of 13C-labeled Ser feeding to the detached shoots of a conifer (Cryptomeria japonica) via the transpiration stream, and compared the labeling patterns of photorespiratory metabolites with those of an angiosperm (Populus nigra). Glycerate was most labeled in P. nigra, whereas glycolate was more labeled than glycerate in C. japonica. The photorespiration pathway involves H2O2-scavenging and H2O2-generating enzymes, catalase (CAT) and glycolate oxidase (GLO), respectively, which are the peroxisomal targeting enzymes in angiosperms. In contrast, database analyses of the peroxisomal targeting signal motifs and analyses of the peroxisomal fractions isolated from C. japonica leaves indicated that the conifer peroxisomes were not a major localization of CAT. These results suggest that the conifer photorespiration pathway has a bypass from Ser to glycolate via the decarboxylation of hydroxypyruvate, because of an imbalance between CAT and GLO activities in peroxisomes. One sentence summaryConifer peroxisome is not a major localization of catalase and yields a unique oxidative decarboxylation path in the photorespiratory pathway.

plant biology↗

bioRxiv2019

Serum miR-379 expression is related to the development and progression of hypercholesterolemia in non-alcoholic fatty liver disease

IntroductionNon-alcoholic fatty liver disease (NAFLD) has a wide spectrum, eventually leading to cirrhosis and hepatic carcinogenesis. We previously reported that a series of microRNAs (miRNAs) mapped in the 14q32.2 maternally imprinted gene region (Dlk1-Dio3 mat) are related to NAFLD development and progression in a mouse model. We examined the suitability of miR-379, a circulating Dlk1-Dio3 mat miRNA, as a human NAFLD biomarker.\n\nMethodsEighty NAFLD patients were recruited for this study. miR-379 was selected from the putative Dlk1-Dio3 mat miRNA cluster because it exhibited the greatest expression difference between NAFLD and non-alcoholic steatohepatitis in our preliminary study. Real-time PCR was used to examine the expression levels of miR-379 and miR-16 as an internal control.\n\nResultsCompared to normal controls, serum miR-379 expression was significantly up-regulated in NAFLD patients. Receiver operating characteristic curve analysis suggested that miR-379 is a suitable marker for discriminating NAFLD patients from controls, with an area under the curve value of 0.72. Serum miR-379 exhibited positive correlations with alkaline phosphatase, total cholesterol, and low-density-lipoprotein cholesterol levels in patients with early stage NAFLD (Brunt fibrosis stage 0 to 1). The correlation between serum miR-379 and cholesterol levels was lost in early stage NAFLD patients treated with statins. Software-based predictions indicated that various energy metabolism-related genes, including insulin-like growth factor-1 (IGF-1) and IGF-1 receptor, are potential targets of miR-379.\n\nConclusionsSerum miR-379 exhibits high potential as a biomarker for NAFLD. miR-379 appears to increase cholesterol lipotoxicity, leading to the development and progression of NAFLD, via interference with the expression of target genes, including those related to the IGF-1 signaling pathway. Our results could facilitate future research into the pathogenesis, diagnosis, and treatment of NAFLD.

molecular biology↗