Masako ASAHINA et al. (2003) International Worm Meeting "A highly conserved yet non-essential coactivator MBF1 may play a role in oxidative stress in C. elegans"

Masako ASAHINA, Marek JINDRA, Susumu Hirose

[International Worm Meeting, 2003]

A small helix-turn-helix protein MBF1 has been maintained throughout the evolution of eukaryotes. Our previous genetic evidence from yeast and Drosophila implicates MBF1 in transcriptional activation mediated by bZIP transcription factors [1,2]. To understand the role of a C. elegans homolog of mbf1, we began to analyze its expression and function. The expression was visualized using an C. elegans mbf1(Cembf1)::GFP reporter and a polyclonal antibody against the Drosophila protein, revealing constitutive and ubiquitous presence of the gene product. Deletions of the transgenic GFP fusion construct showed that a relatively short (223 bp) upstream region was sufficient for Cembf1 expression. Functional conservation of mbf1 was tested in yeast (S. cerevisiae) mbf1 disruptant that is viable but sensitive to histidine starvation [1]. The growth of the mbf1 mutant yeast was partially restored by transformation with the C. elegans mbf1 homolog, indicating that the mechanism of MBF1 action could be conserved from unicellular to multicellular organisms. RNA interference silencing of mbf1 in C. elegans showed fairly wild type phenotype although the Cembf1 dsRNA effectively suppressed GFP in the Cembf1::GFP transgenic worms. This suggests that, as in yeast and flies, mbf1 is not an essential gene in C. elegans. The question then arises of what makes mbf1 important enough to be conserved during evolution. Our preliminary data show that the mbf1 RNAi treated worms are more sensitive to hydrogen peroxide, implying that MBF1 is required to withstand oxidative stress. A microarray analysis is underway to find what genes might be affected in mbf1 RNAi background during the hydrogen peroxide challenge. [1] Takemaru K, Harashima S, Ueda H and Hirose S. (1998) Mol. Cell Biol. 18, 4971-76 [2] Liu QX, Jindra M, Ueda H, Hiromi Y, Hirose S. (2003) Development 130, 719-28

Toshihiro Sassa et al. (2001) International C. elegans Meeting "THE C.ELEGANS PROTEIN PHOSPHATASE 1 HOMOLOGUE(CEPP1) IS ESSENTIAL FOR EMBRYOGENESIS AND POST-EMBRYONIC DEVELOPMENT"

Ryuji Hosono, Keiko Gengyo-Ando, Shohei MItani, Shin-ichi Harada, Keiichiro Kitamura, Kanako Hirano, Toshihiro Sassa, Hiroko Ueda

[International C. elegans Meeting, 2001]

Protein dephosphorylation functions as modulator in many biological events. To elucidate the regulatory aspects of protein dephosphorylation, we identified and cloned the gene encoding protein phosphatase 1 homologue( Ce PP1), which was found within cosmid F56C9 located LGIII. From the expression pattern of the Ce PP1::GFP fusion protein and in situ hybridization, the gene is ubiquitously but especially strongly expressed in gonad, neuron, muscle and intestine. Microinjection of double stranded RNA of this gene induced an embryonic arrest at multicellular stages. To characterize in more detail the function of gene( pp1-1 ) encoding Ce PP1, we isolated a mutant( tm291 ) by TMP/UV method, which had a deletion of exon VII of pp1-1 . Hermaphrodites heterozygous for the pp1-1 mutation produce homozygous embryos which can hatch and grow to adulthood. However, progeny produced from hermaphrodites homozygous for its mutation almost (96%) die at multicellular stages during embryogenesis. DAPI staining of embryos revealed that the presence of chromatin bridge and multinucleated cells, suggesting the gene being implicated in chromosome condensation and segregation. However, the staining of the mutant with antisera against alpha-tubulin was normal. These observations are reminiscent of the air-1 /aurora-related kinase gene defective mutants(Schumacher et al ., Dev. 125 4391, 1998) or nuclear lamin( lmn-1 ) defective embryos(RNAi)(Liu et al ., Mol.Biol.Cell 11 3937, 2000). The defect of karyokinesis in the pp1-1 mutants may be elucidated by the following results; Ce PP1 expressed in HEK 293 cells catalyzes Pi release from phospho-histone H3 phosphorylated by PKA. The morphological abnormality of homozygous mutant at the post-embryonic stage is also observed including vulva and gonad formation. These phenotypes are also reminiscent of stu-7/air-2 defective mutants (Woollard and Hodgkin, Mech. Dev., 82 95, 1999). The pp1-1 gene was strongly expressed in the AFD and ASE amphidal neurons. We therefore are ongoing to elucidate the implication of the gene in both neurons. As mutant showed normal movement, we can examine chemotactic and thermotactic behavior which were sensed by ASE and AFD, respectively. Preliminary studies showed that the mutants displayed normal attraction to NaCl, but abnormal thermotactic behavior.

Lin Lin et al. (2006) East Asia C. elegans Meeting "Mutants Having Altered Preference of Chemotaxis in Simultaneous Presentation of Two Attractants"

Tarou OGURUSU, Ryuzo SHINGAI, Tsutomu SATO, Tokumitsu WAKABAYASHI, Lin Lin, Tomohiro OIKAWA

[East Asia C. elegans Meeting, 2006]

Mutants Having AlteredPreference of Chemotaxis in Simultaneous Presentation of Two Attractants Lin Lin, <SUP>a</SUP> Tokumitsu WAKABAYASHI, <SUP>b</SUP> Tomohiro OIKAWA,<SUP>a</SUP>Tsutomu SATO,<SUP>b</SUP> Tarou OGURUSU <SUP>a, b</SUP> and Ryuzo SHINGAI<SUP>a,b,</SUP> <SUP>a</SUP>Department of Computer and Information Science, Graduate School of Engineering, Iwate University, 4-3-5 Ueda, Morioka, 020-8551 Japan <SUP>b</SUP>Laboratory of Bioscience, Faculty of Engineering, Iwate University, 4-3-5 Ueda, Morioka, 020-8551 Japan Upon presentation of two distinct chemoattractants such as sodium acetate and diacetyl simultaneously, the worms were preferentially attracted by one of these chemoattractants. We isolated two mutants having altered preference of chemotaxis behavior toward simultaneous presentation of sodium acetate and diacetyl. chep-1(qr1) (for chemosensory preference) mutant preferred sodium acetate to diacetyl, while chep-2(qr2) mutant preferred diacetyl to sodium acetate in simultaneous presentation of these chemoattractants. chep-1(qr1) showed decreased preference toward diacetyl in the simultaneous two-spot presentation in all conditions tested. The decreased preference toward diacetyl could be explained by the sole effect of the defective chemotaxis toward diacetyl. The chemotaxis behavior of chep-2(qr2) mutant in simultaneous presentation suggested the function of chep-2 gene products within the chemosensory informational integration pathway as well as in the chemosensory pathway. Sodium acetate is detected by the ASEL chemosensory neuron and the neuron sends its synaptic output onto AIA, AIB, AIY interneurons and several other sensory neurons. A low concentration of diacetyl is detected by a bilateral pair of AWA sensory neurons and send their synaptic outputs onto AIY and AIZ neurons and several sensory neurons including ASE. Multiple information detected separately by ASEL and AWA neurons could be integrated directly within the ASEL neuron and within the AIY interneurons. The integration within the AIA, AIB and/or AIZ interneurons were also possible. chep-2 gene product may function in one to several of these neurons to regulate the informational integration pathway.