Tsuyama, Kenji, Takabayashi, Yusuke, Maruyama, Rika, Haruta, Nami, Sugimoto, Asako, Kubota, Yukihiko
[
International Worm Meeting,
2013]
During morphogenesis, cells undergo dynamic changes including polarization, migration and rearrangement of cell groups. To identify new genes involved in epidermal morphogenesis, we performed an RNAi screen of ~800 embryonic lethal genes by live imaging, and identified B0464.2 whose depletion caused highly penetrant defects in body elongation in late embryogenesis. B0464.2 encodes the C. elegans ortholog of Ctr9 (Ce-Ctr9), a component of the Polymerase-Associated Factor 1 (PAF1) complex. The PAF1 complex consists of five conserved proteins (Paf1, Ctr9, Cdc73, Leo1 and Rtf1), and is implicated in diverse transcription related processes including transcriptional elongation, 3'-terminal end processing, and histone modification. We found by RNAi that, in addition to the Ce-Ctr9, orthologs of all other four components of the PAF1 complex were required for epidermal morphogenesis. Time-lapse analyses by DIC and with DLG-1::GFP (an epidermal junction maker), mCherry::TBB-2 (a microtubule marker) and VAB-10(ABD)::mCherry (an F-actin marker) revealed that, while depletion of components of the PAF1 complex did not affect the number of epithelial cells, it affected cell shape changes and cell positioning during epidermal enclosure and body elongation. Abnormalities in muscles and neurons were not detected in these embryos. Genomic translational GFP/mCherry-fusion constructs of Ce-Paf1, Ce-Leo1 and Ce-Rtf1 revealed that these proteins were localized to the nuclei of virtually all embryonic cells. They appeared not tightly associated with chromatins, because these proteins were diffused to the cytoplasm during NEBD. A deletion mutant of Ce-Leo1 exhibited maternal effect embryonic lethality with epidermal defects similar to the RNAi embryos, which were rescued by mCherry::Leo1 expression under an epidermis-specific promoter. These results indicate that the PAF1 complex regulates the epidermal cell shape change and migration during embryonic morphogenesis in a cell-autonomous manner.
Kanzaki, Natsumi, Hoshi, Yuki, Kumagai, Ryohei, Sugimoto, Asako, Kikuchi, Taisei, Namai, Satoshi, Tsuyama, Kenji
[
International Worm Meeting,
2017]
Caenorhabditis sp. 34 is a sister species of C. elegans recently isolated from the syconia of the fig Ficus septica on Ishigaki Island, Japan (see abstract by T. Kikuchi, et al.). C. sp. 34 is gonochoric and shares typological key characters with other Elegans supergroup species, but strikingly, adults are nearly twice as long as C. elegans. The optimal culture temperature for C. sp. 34 is significantly higher (27 deg C) than that of C. elegans (20 deg C). Young adult males and females tend to form clumps, and Dauer larvae are rarely observed in laboratory culture conditions. Recently the C. sp. 34 genome assembly was produced into six chromosomes (see abstract by T. Kikuchi, et al.). The marked differences from C. elegans in morphology, behaviors and ecology, and the availability of the complete genome sequence make C. sp. 34 highly attractive for comparative and evolutionary studies. To make C. sp. 34 genetically tractable, we have been developing genetic and molecular techniques and tools. Stable transgenic lines of C. sp.34 could be obtained by microinjecting marker plasmids commonly used in C. elegans, although the efficiency was lower than that in C. elegans. Both soaking and feeding RNAi was as effective as in C. elegans. A panel of antibodies against C. elegans proteins successfully recognized expected structures in C. sp. 34 by immunofluorescence. Thus, many of the rich genetic and molecular resources for C. elegans can be directly used for C. sp. 34 studies. We well present some of the comparative analyses of gene functions regarding the body size, germ cell formation and sex determination.