Yoshida K et al. (2012) Nat Commun "Odour concentration-dependent olfactory preference change in C. elegans."

Ishihara T, Tagawa T, Oda S, Wakabayashi T, Hirotsu T, Iino Y, Yoshida K

[Nat Commun, 2012]

The same odorant can induce attractive or repulsive responses depending on its concentration in various animals including humans. However, little is understood about the neuronal basis of this behavioural phenomenon. Here we show that Caenorhabditis elegans avoids high concentrations of odorants that are attractive at low concentrations. Behavioural analyses and computer simulation reveal that the odour concentration-dependent behaviour is primarily generated by klinokinesis, a behavioural strategy in C. elegans. Genetic analyses and lesion experiments show that distinct combinations of sensory neurons function at different concentrations of the odorant; AWC and ASH sensory neurons have critical roles for attraction to or avoidance of the odorant, respectively. Moreover, we found that AWC neurons respond to only lower concentrations of the odorant, whereas ASH neurons respond to only higher concentrations of odorant. Hence, our study suggests that odour concentration coding in C. elegans mostly conforms to the labelled-line principle where distinct neurons respond to distinct stimuli.

Yoshida, K. et al. (2019) International Worm Meeting "Identification of a novel RNAi-related factor that promotes intercellular transport of double-stranded RNA."

Yoshida, K., Mitani, S., Yoshina, S., Suehiro, Y.

[International Worm Meeting, 2019]

During the process of systemic RNA interference (RNAi), consisting of secretion from donner cells and uptake into recipient cells of double-stranded RNA (dsRNA), membrane trafficking is thought to play important roles. However, it is not well understood which membrane trafficking pathways are involved in the transport of dsRNA. To identify the secretory pathway of dsRNA in systemic RNAi, we screened for mutants that show defects in RNAi triggered by ingested dsRNA under the genetic background of double mutant for rab-3 and aex-6/rab-27, which encode Rab-GTPases regulating the trafficking pathway for secretion. We identified two independent mutations in the Y39A3CL.1 gene that are partially deficient in the feeding RNAi. Another mutant carrying a Y39A3CL.1 deletion also showed RNAi resistance both in somatic cells and in the germline. Y39A3CL.1 did not genetically interact with rab-3 and aex-6 double mutation whereas it was originally identified by mutagenizing the double mutant. Y39A3CL.1 encodes a protein with no known domains and has no homologs outside of nematodes. Y39A3CL.1 is expressed in various tissue including hypodermis, pharynx and intestine. Tissue-specific rescue experiments revealed that Y39A3CL.1 can act in a cell non-autonomous manner during systemic RNAi. We examined subcellular localization of Y39A3CL.1 fused with RFP and found that it co-localizes with late endosome and lysosome markers. These results suggest that Y39A3CL.1 is a novel RNAi-related factor which may act in an endomembrane-trafficking pathway and promote intercellular transport of dsRNA.

Yoshida K et al. (2023) iScience "Distinct pathways for export of silencing RNA in Caenorhabditis elegans systemic RNAi."

Yoshina S, Dejima K, Yoshida K, Mitani S, Suehiro Y

[iScience, 2023]

Dietary supplied double-stranded RNA (dsRNA) can trigger RNA interference (RNAi) systemically in some animals, including the nematode <i>Caenorhabditis elegans</i>. Although this phenomenon has been utilized as a major tool for gene silencing in <i>C.&#xa0;elegans</i>, how cells spread the silencing RNA throughout the organism is largely unknown. Here, we identify two novel systemic RNAi-related factors, REXD-1 and TBC-3, and show that these two factors together with SID-5 act redundantly to promote systemic spreading of dsRNA. Animals that are defective in all REXD-1, TBC-3, and SID-5 functions show strong deficiency in export of dsRNA from intestinal cells, whereas cellular uptake and processing of dsRNA and general secretion events other than dsRNA secretion are still functional in the triple mutant animals. Our findings reveal pathways that specifically regulate the export of dsRNA in parallel, implying the importance of spreading RNA molecules for intercellular communication in organisms.

Yoshida K et al. (2024) STAR Protoc "Protocol for forward genetic screening to identify novel factors involved in a biological process in Caenorhabditis elegans."

Yoshida K, Suehiro Y, Mitani S

[STAR Protoc, 2024]

Forward genetic screens have been a powerful tool for discovering genes involved in various biological processes in Caenorhabditis elegans. Here, we present a protocol for forward genetic screening to identify novel factors involved in a biological process in C.elegans. We describe steps for mutagenesis, screening, and backcrossing. To save time and effort, we also detail procedures for utilizing whole-genome sequencing to exclude mutants of previously characterized genes from crosses for mapping mutations. For complete details on the use and execution of this protocol, please refer to Yoshida et&#xa0;al.¹.

Yoshida, K. et al. (2011) International Worm Meeting "Olfactory preference switch depending on odor concentration is mediated by combinatorial change of acting sensory neurons."

Hirotsu, T., Oda, S., Tagawa, T., Yoshida, K., Ishihara, T., Iino, Y.

[International Worm Meeting, 2011]

Many organisms exhibit olfactory preference towards most odorants. It is empirically known that the perceived quality of an odorant changes depending on its concentration in some organisms including humans. However, the neuronal basis of the preference change depending on odor concentration is still elusive. Here we show C. elegans also shows changes of olfactory preference depending on odor concentration. When concentrations of various attractive odorants such as isoamyl alcohol are increased, animals become to show avoidance of them. Behavioral analyses and computer simulation revealed that the behavioral change depending on concentration of an odorant is mainly mediated by one of two known behavioral strategies, klinokinesis (the pirouette mechanism). Namely, the dependence of the frequency of pirouette behavior on temporal changes in odor concentration is reversed in different concentration ranges. On the other hand, the other behavioral strategy klinotaxis (the weathervane mechanism) is employed in the direction to drive attraction to the odorant regardless of its concentration. We next examined what molecules mediate these behaviors, and found that ODR-3 G protein a subunit mediates klinokinesis and klinotaxis in the concentration-dependent chemotaxis to isoamyl alcohol. Moreover, ODR-3 transmits the olfactory signaling depending on isoamyl alcohol concentration by acting in different sensory neurons. We further show that distinct combinations of sensory neurons function at different concentrations of isoamyl alcohol: AWC sensory neurons are known to mediate attraction to lower concentration of isoamyl alcohol, whereas ASH, AWB and ADL sensory neurons are required for avoidance of higher concentration of it. Calcium imaging revealed that activity patterns of these sensory neurons dramatically change depending on isoamyl alcohol concentration: the higher concentration of isoamyl alcohol fails to elicit responses of AWC, which strongly responds to lower concentration of the odorant, and elicits particular activity patterns of AWB and ASH. Hence, our study shows that the information processing which regulates olfactory preference change depending on odor concentration is clearly explained by the labeled-line theory.

Yoshida K et al. (2010) Neuronal Development, Synaptic Function and Behavior, Madison, WI "Neural and Behavioral Mechanisms of Response to an Odorant Depending on its Concentration"

Ishihara T, Yoshida K, Tagawa T, Iino Y, Oda S, Hirotsu T

[Neuronal Development, Synaptic Function and Behavior, Madison, WI, 2010]

The perceived quality of an odorant can differ with a change in its concentration in many species. In C. elegans, it was reported that several odorants such as benzaldehyde and 2,4,5-trimethylthiazole, act as attractants at low concentrations, but as repellents at high concentrations. We found that this dose-dependent behavioral change was also observed in chemotaxis to all the odorants tested (isoamyl alcohol, diacetyl, and 2,3-pentanedione). To examine the underlying neural circuit, we performed laser ablation of individual neurons and found that avoidance of high concentration of isoamyl alcohol is regulated by multiple sensory neurons. Moreover, we identified some interneurons which positively mediate this avoidance. It was previously reported that AWC sensory neurons regulate attraction to diluted isoamyl alcohol and are activated by the removal of isoamyl alcohol. Our results of calcium imaging revealed that AWB and ASH sensory neurons, which are both known to mediate an avoidance behavior, respond to the removal and addition of isoamyl alcohol, respectively. Recently, we reported that worms employ two strategies (the pirouette and weathervane strategies) for efficient chemotaxis. To investigate the behavioral mechanisms of avoidance of isoamyl alcohol at the high concentration, we analyzed this behavior by using multi-worm tracking system. Though the pirouette mechanism was mainly used to regulate this avoidance behavior, interestingly, the weathervane mechanism was employed to curve toward isoamyl alcohol. Computer simulation showed that behaviors of model animals using both strategies are quantitatively similar to those of real worms. We also looked for molecules involved in avoidance of isoamyl alcohol at the high concentration, and found that mutants of odr-3, which encodes a G-protein alpha subunit, show a severe defect in this avoidance. Furthermore, cell-specific rescue experiments suggested that the function of ODR-3 in AWB sensory neurons is important for avoidance of isoamyl alcohol.

Iino Y et al. (2010) Tanpakushitsu Kakusan Koso "[How animals reach their destination?: Two strategies of chemotaxis in C. elegans]"

Yoshida K, Iino Y

[Tanpakushitsu Kakusan Koso, 2010]

Iino Y et al. (2009) J Neurosci "Parallel use of two behavioral mechanisms for chemotaxis in Caenorhabditis elegans."

Iino Y, Yoshida K

[J Neurosci, 2009]

Caenorhabditis elegans shows chemotaxis to various odorants and water-soluble chemoattractants such as NaCl. Previous studies described the pirouette mechanism for chemotaxis, in which C. elegans quickly changes the direction of locomotion by using a set of stereotyped behaviors, a pirouette, in response to a decrease in the concentration of the chemical. Here, we report the discovery of a second mechanism for chemotaxis, called the weathervane mechanism. In this strategy animals respond to a spatial gradient of chemoattractant and gradually curve toward higher concentration of the chemical. By computer simulation, we find that both of these mechanisms contribute to chemotaxis and both mechanisms need to act in parallel for efficient chemotaxis. Using laser ablation of individual neurons to examine the underlying neural circuit, we find the ASE sensory neurons and AIZ interneurons are essential for both the pirouette and weathervane mechanisms in chemotaxis to NaCl. Salt-conditioned animals show reversed responses in both of these behaviors, leading to avoidance of NaCl. These results provide a platform for detailed molecular and cellular analyses of chemotaxis and its plasticity in this model organism.

Yoshida S et al. (2019) FEBS Lett "Multiple functions of DYRK2 in cancer and tissue development."

Yoshida K, Yoshida S

[FEBS Lett, 2019]

Dual-specificity tyrosine-regulated kinases (DYRKs) are evolutionarily conserved from yeast to mammals. Accumulating studies have revealed that DYRKs have important roles in regulation of the cell cycle and survival. DYRK2, a member of the class II DYRK family protein, is a key regulator of p53, and phosphorylates it at Ser46 to induce apoptosis in response to DNA damage. Moreover, recent studies have uncovered that DYRK2 regulates G1/S transition, epithelial-mesenchymal-transition (EMT), and stemness in human cancer cells. DYRK2 also appears to have roles in tissue development in lower eukaryotes. Thus, the elucidation of mechanisms for DYRK2 during mammalian tissue development will promote the understanding of cell differentiation, tissue homeostasis, and congenital diseases as well as cancer. In this review, we discuss the roles of DYRK2 in tumor cells. Moreover, we focus on DYRK2-dependent developmental mechanisms in several species including fly (Drosophila), worm (C. elegans), zebrafish (Danio rerio), and mammals.

Yoshida A et al. (2015) Genes Brain Behav "A glial K(+) /Cl(-) cotransporter modifies temperature-evoked dynamics in C. elegans sensory neurons."

Suzuki T, Ihara K, Mori I, Nakano S, Higashiyama T, Yoshida A

[Genes Brain Behav, 2015]

K(+) /Cl(-) cotransporters (KCCs) are known to be crucial in the control of neuronal electrochemical Cl(-) gradient. However, the role of these proteins in glial cells remains largely unexplored despite a number of studies showing expression of KCC proteins in glial cells of many species. Here, we show that the C. elegans K(+) /Cl(-) cotransporter KCC-3 is expressed in glial-like cells and regulates the thermosensory behavior through modifying temperature-evoked activity of a thermosensory neuron. Mutations in the kcc-3 gene were isolated from a genetic screen for mutants defective in thermotaxis. KCC-3 is expressed and functions in the amphid sheath glia that ensheathes the AFD neuron, a major thermosensory neuron known to be required for thermotaxis. A genetic analysis indicated that the regulation of the thermosensory behavior by KCC-3 is mediated though AFD, and we further show that KCC-3 in the amphid sheath glia regulates the dynamics of the AFD activity. Our results reveal a novel mechanism by which the glial KCC-3 protein non-cell autonomously modifies the stimulus-evoked activity of a sensory neuron and highlight the functional importance of glial KCC proteins in modulating the dynamics of a neural circuitry to control an animal behavior.