Shimada A et al. (2010) Z Naturforsch C "Nematicidal activity of beauvericin produced by the fungus Fusarium bulbicola."

Koshino H, Kimura Y, Shimada A, Fujioka S

[Z Naturforsch C, 2010]

A nematicide, beauvericin (1), was isolated from cultures of the fungus Fusarium bulbicola, and its structure was identified by spectroscopic analysis. Compound 1 showed nematicidal activities against the pine wood nematode Bursaphelenchus xylophilus and the free-living nematode Caenorhabditis elegans.

Shimada H et al. (2003) Trace Elements and Electrolytes "Metallothionein gene expression in the larvae of Caenorhabditis elegans is a potential biomarker for cadmium and mercury."

Ishibashi H, Shimada H, Kohra S, Tominaga N, Ura K, Mitsui Y, Arizono K

[Trace Elements and Electrolytes, 2003]

Previously, we reported that the induction of vitellogenin mRNA in the larvae of Caenorhabditis elegans (C. elegans) can be used as a biomarker for short-term screening of environmental endocrine disrupters. Therefore, the present study was designed to determine if the induction of metallothionein (MT) mRNA in the larvae of C. elegans would be a biomarker for short-term screening of heavy metals. The larvae were exposed to various concentrations of cadmium (Cd), mercury (Hg), zinc chloride (Zn), cupper chloride (Cu) or lead acetate (Pb) for 2h. Cd (1, 10 and 100 uM) and Hg (0.01, 0.1, 1 and 10 uM) exposures resulted in the marked induction of MT-I and MT-II mRNAs in the larvae of C. elegans as measured by reverse transcription polymerase chain reaction. The Cd and Hg induction of MT-II was higher than that of MT-I mRNA, and concentration-dependent increase was observed in MT-II but not in MT-I. Time course analysis for MT-I and MT-II expressions with Cd and Hg were also determined. Cd induction of MT-I and MT-II mRNAs reached a peak at 2 h after the exposure (10 uM), and the levels of MT-II were higher than that of MT-I. For Hg, an initial peak of induction of MT-II mRNA occurred 15 minutes after the exposure (0.1 uM), and the levels reached maximum by 2 h. The initial peak of induction of MT-I mRNA occurred much later (about 2 h after Hg exposure) than MT-II mRNA. These results indicate that the induction of MT-II mRNA in the larvae of C. elegans can be used as a potential biomarker

Shimada M et al. (2006) Mol Biol Cell "Proteasomal ubiquitin receptor RPN-10 controls sex determination in Caenorhabditis elegans."

Yokosawa H, Shimada M, Tanaka K, Kanematsu K, Kawahara H

[Mol Biol Cell, 2006]

Monitoring Editor: Thomas Sommer The ubiquitin-binding RPN-10 protein serves as a ubiquitin receptor that delivers client proteins to the 26S proteasome. Although ubiquitin recognition is an essential step for proteasomal destruction, deletion of the rpn-10 gene in yeast does not influence viability, indicating redundancy of the substrate delivery pathway. However, their specificity and biological relevance in higher eukaryotes is still enigmatic. We report herein that knockdown of the rpn-10 gene, but not any other proteasome subunit genes, sexually transforms hermaphrodites to females by eliminating hermaphrodite spermatogenesis in Caenorhabditis elegans. The feminization phenotype induced by deletion of the rpn-10 gene was rescued by knockdown of tra-2, one of sexual fate decision genes promoting female development, and its downstream target tra-1, indicating that the TRA-2-mediated sex-determination pathway is crucial for the Deltarpn-10-induced sterile phenotype. Intriguingly, we found that coknockdown of rpn-10 and functionally related ubiquitin ligase ufd-2 overcomes the germline musculinizing effect of fem-3(gf). Furthermore, TRA-2 proteins accumulated in rpn-10-defective worms. Our results show that the RPN-10-mediated ubiquitin pathway is indispensable for control of the TRA-2-mediated sex-determining pathway.

Shimada M et al. (2006) Genes Cells "OMA-1 is a P granules-associated protein that is required for germline specification in Caenorhabditis elegans embryos."

Shimada M, Yokosawa H, Kawahara H

[Genes Cells, 2006]

In Caenorhabditis elegans, CCCH-type zinc-finger proteins have been shown to be involved in the differentiation of germ cells during embryonic development. Previously, we and others have identified novel redundant CCCH-type zinc-finger proteins, OMA-1 and OMA-2, that are involved in oocyte maturation. In this study, we report that the cytoplasmic expression level of OMA-1 protein was largely reduced after fertilization. In contrast to its cytoplasmic degradation, OMA-1 was found to accumulate exclusively on P granules in germline blastomeres during embryogenesis. A notable finding is that embryos with partially suppressed oma-1; oma-2 expression showed inappropriate germline specification, including abnormal distributions of PGL-1, MEX-1 and PIE-1 proteins. Thus, our results suggest that oma gene products are novel multifunctional proteins that participate in crucial processes for germline specification during embryonic development.

Shimada Y et al. (2023) Biomolecules "Spermiogenesis in Caenorhabditis elegans: An Excellent Model to Explore the Molecular Basis for Sperm Activation."

Kanazawa-Takino N, Nishimura H, Shimada Y

[Biomolecules, 2023]

<i>C. elegans</i> spermiogenesis converts non-motile spermatids into motile, fertilization-competent spermatozoa. Two major events include the building of a pseudopod required for motility and fusion of membranous organelles (MOs)-intracellular secretory vesicles-with the spermatid plasma membrane required for the proper distribution of sperm molecules in mature spermatozoa. The mouse sperm acrosome reaction-a sperm activation event occurring during capacitation-is similar to MO fusion in terms of cytological features and biological significance. Moreover, <i>C. elegans fer-1</i> and mouse <i>Fer1l5</i>, both encoding members of the ferlin family, are indispensable for MO fusion and acrosome reaction, respectively. Genetics-based studies have identified many <i>C. elegans</i> genes involved in spermiogenesis pathways; however, it is unclear whether mouse orthologs of these genes are involved in the acrosome reaction. One significant advantage of using <i>C. elegans</i> for studying sperm activation is the availability of in vitro spermiogenesis, which enables combining pharmacology and genetics for the assay. If certain drugs can activate both <i>C. elegans</i> and mouse spermatozoa, these drugs would be useful probes to explore the mechanism underlying sperm activation in these two species. By analyzing <i>C. elegans</i> mutants whose spermatids are insensitive to the drugs, genes functionally relevant to the drugs' effects can be identified.

Masumi Shimada et al. (2005) International Worm Meeting "Ubiquitin-binding subunit of 26S proteasome is essential for TRA-2-mediated sex determination in Caenorhabditis elegans"

Hiroyuki Kawahara, Masumi Shimada

[International Worm Meeting, 2005]

RPN-10 subunit of 26S proteasome as well as several UBA domain proteins can bind to a polyubiquitin chain and are thought to play a role as ubiquitin receptors of 26S proteasome. Although it was thought that substrate recognition is an essential step in the proteasome-mediated protein degradation, deletion of RPN-10 and UBA-domain protein genes in yeast does not influence the viability of cells and instead causes only a mild phenotype, suggesting that they are redundantly involved in substrate delivery to the proteasome, though their functional difference is still enigmatic. In this study, we report that suppression of RPN-10 did not cause a lethal phenotype in C. elegans, as is the case in yeast. In accordance with this, there are few defects in 26S proteasome architecture in rpn-10 (RNAi) C. elegans, though polyubiquitinated proteins accumulated in these worms. Although RPN-10 is dispensable for viability, we found that RPN-10 is required for appropriate sex determination in C. elegans. Suppression of the RPN-10 sexually transforms F1 hermaphrodites to females by eliminating hermaphrodite spermatogenesis. The feminization phenotype induced by rpn-10 (RNAi) is rescued by co-suppression of tra-2 and its downstream target tra-1, indicating that TRA-2-mediated inhibitory pathway is crucial for rpn-10 (RNAi)-induced sterile phenotype. We found that TRA-2 proteins, either ubiquitinated forms or fragments of TRA-2 ICD, accumulate in rpn-10 (RNAi) worms, though the amount of tra-2 transcript is not influenced. Thus, it can be explained that suppression of RPN-10 induces a sex determination defect due to specific failure of degradation on polyubiquitinated TRA-2 protein and resulting overproduction of its intracellular domain. Our results show that the RPN-10-mediated ubiquitin-dependent pathway is indispensable for the control of the TRA-2-mediated sex determining signaling pathway. We speculate that accumulation of TRA-2 is an almost exclusive defect in RPN-10-suppressed conditions, except for the secondary defects in OMA-2 aggregation in arrested oocytes. We believe that the most of the ubiquitinated substrates are normally metabolized in rpn-10 (RNAi) worms by a redundant pathway to RPN-10 recognition, such as the pathway mediated by the UBA-domain proteins RAD-23 and DSK-2. Based on these observations, we propose that the recruitment of substrates to the proteasome by ubiquitin receptors provides an additional layer of substrate selection in the ubiquitin-proteasome pathway, and supports the notion that RPN-10 functions as a multiubiquitin receptor for specific substrate proteins and that no other protein is involved in this function. Kawahara, H. et al. (2000) EMBO J. 19, 4144-4153.

Masumi Shimada et al. (2006) Development & Evolution Meeting "Analyses of Endogenous TRA-2 Protein using Newly Established anti-TRA-2 ICD Antibody"

Hiroyuki Kawahara, Satomi Harukuni, Hideyoshi Yokosawa, Masumi Shimada

[Development & Evolution Meeting, 2006]

The genetic basis of sex determination has been analyzed extensively in the nematode, which has two naturally occurring sexes, an XX hermaphrodite and an XO male. Most of the sexual fate decision genes have been identified and have been shown to function in a cascade of negative regulation. Sex determination requires post-transcriptional regulation of a key gene, tra-2. Genetic studies have shown that tra-2 promotes female development since loss-of-function mutations of tra-2 transform XX animals into pseudomales that do not produce oocytes. TRA-2 is thought to be a large transmembrane protein, and is controlled by its ligand HER-1, by proteolytic cleavage of intracellular domain (ICD), by alternative splicing of mRNA, by nuclear export of mRNA and by translational regulation through 3'UTR-binding proteins. However, the cellular behavior of endogenous TRA-2 protein is still enigmatic as a high titer antibody had not been available. To detect the endogenous TRA-2 protein directly, we recently established an antiserum to the 7th intracellular domain of TRA-2 (TRA-2 ICD). With this antibody, we found the nuclei of hermaphrodite intestine, a female somatic tissue, were clearly immunostained. We believe that the staining is specific to endogenous TRA-2 protein, as our tra-2(RNAi) on fem-3 (e1996lf) greatly reduced the intensity of nuclear staining. Furthermore, we confirmed that nuclei of tra-2(e2020gf) were stained much stronger than that of wild-type, while there are essentially no nuclear signals in tra-2(e1095lf) or in wild-type males. Lum et al. (2000) reported that GFP-TRA-2c, overproduced from the heat shock promoter, localized predominantly to nuclei in some somatic cells, and caused completely somatic feminization of XO animals. Our results supporting the possibility that endogenous TRA-2 (ICD) and nuclear transcriptional regulator TRA-1 have the opportunity to interact in vivo. With this antibody, we are now trying to examine the TRA-2 distribution in C. elegans germline during the course of development. Our results will be discussed in the meeting. We thank Dr. S. Mitani, Dr. K. Gengyo-Ando, Dr. T. Schedl, Dr. B.Grant and the Caenorhabditis Genetics Center (CGC) for providing the mutant strains. We also thank Dr. H. Sorimachi,Dr. I. Kawasaki, Drs. Ken and Miyuki Sato and many other people in C. elegans society for helpful suggestions and discussions.

Masumi Shimada et al. (2003) International Worm Meeting "Possible roles of MOE-1/OMA-1 CCCH-type zinc-finger family proteins in C. elegans oogenesis and embryogenesis"

Hiroyuki Kawahara, Masumi Shimada

[International Worm Meeting, 2003]

Oocyte maturation is an important prerequisite for the production of progeny. Although several germ-line mutations have been reported, the precise mechanism by which the last step of oocyte maturation is controlled remains unclear. In Caenorhabditis elegans, CCCH-type zinc-finger proteins have been shown to be involved in germ cell formation, although their involvement in oocyte maturation had not been fully investigated. Using a multiple RNAi technique, we have identified three redundant CCCH-type zinc-finger genes, named by us moe-1, -2 and moe-3, as a group related by functions and nucleotide sequence1. Similar results were reported by Detwiler et al.(2001), and they gave the names MOE-1 and MOE-2 to OMA-1 and OMA-2, respectively2. We and Detwiler et al. have found that they have overlapping functions that are crucial for oocyte maturation; i.e. simultaneous removal of these proteins by RNAi induces arrest and expansion of growing oocytes. The results of our in situ hybridization have revealed that each of the moe/oma transcripts is expressed from the distal to proximal region of the gonad, while their corresponding proteins accumulate specifically in the cytoplasm of growing oocytes as well as on P granules. Thus, these gene products participate in processes in the final step of the meiotic cell cycle control, a novel function for CCCH-type zinc-finger family proteins thus far discovered. Although MOE-2/OMA-2 protein is rapidly removed from P granules after fertilization, we found that MOE-2/OMA-2 associated with the centrosome-peripheral structure in dividing blastomeres. Our results suggest that moe/oma gene products are unique multifunctional proteins in terms of their redundancy and characteristic behavior during the course of oocyte maturation and subsequent embryogenesis. 1 Shimada, M., Kawahara, H., and Doi, H. (2002). Genes Cells 7, 933-947. 2 Detwiler, M.R., Reuben, M., Li, X., Rogers, E., and Lin, R. (2001). Dev. Cell 1, 187-199.

Shimada M et al. (2002) Genes Cells "Novel family of CCCH-type zinc-finger proteins, MOE-1, -2 and -3, participates in C. elegans oocyte maturation."

Shimada M, Doi H, Kawahara H

[Genes Cells, 2002]

Background: Oocyte maturation is an important prerequisite for the production of progeny. Although several germ-line mutations have been reported, the precise mechanism by which the last step of oocyte maturation is controlled remains unclear. In Caenorhabditis elegans , CCCH-type zinc-finger proteins have been shown to be involved in germ cell formation, although their involvement in oocyte maturation has not been fully investigated. Results: Using a multiple RNAi technique, we have identified three novel redundant CCCH-type zinc-finger genes, named by us moe-1 , -2 (oma-1 , -2 ) and moe-3 , as a group related by functions and nucleotide sequence. Although a single RNAi of each moe gene was not effective, double or triple RNAi induced defects in oocyte maturation. We found that each moe transcript was expressed from the distal to proximal region of the gonad, while their corresponding proteins are accumulated exclusively in proximal oocytes, with a close association to germ granules. Although MOE-2 protein is rapidly removed from germ granules after fertilization, we found that MOE-2 associates with the centrosome-peripheral structure in dividing blastomeres. Conclusions: Our results suggest that moe gene products are unique multifunctional proteins in terms of their redundancy and characteristic behaviour during the course of oocyte maturation. These gene products participate in processes in the final step of the meiotic cell cycle control, a novel function for CCCH-type zinc-finger family proteins thus far discovered.

Ohtani K et al. Z Naturforsch C "Nematicidal activities of 4-hydroxyphenylacetic acid and oidiolactone D produced by the fungus Oidiodendron sp."

Ohtani K, Kawano T, Kimura Y, Shimada A, Fujioka S

[Z Naturforsch C]

Two nematicides, 4-hydroxyphenylacetic acid (4-HPA) (1) and oidiolactone D (2), were isolated from cultures of the fungus Oidiodendron sp., and their structures were identified by spectroscopic analyses. Compound 2 showed nematicidal activities against the root-lesion nematode, Pratylenchus penetrans, and the pine wood nematode, Bursaphelenchus xylophilus. Compound 1 was also active against these two nematodes but to a lesser extent.