Natesan, Balasubramanian (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "Entomopathogenic nematode Steinernema carpocapsae secreted protease role on host immune suppression"

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C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany,

2010]

Entomopathogenic nematode Steinernema carpocapsae secreted protease role on host immune suppression Natesan Bala subramanian, Duarte Toubarro and Nelson Simes CIRN and Department of Biology, University of Azores , 9500-801 Ponta Delgada , Portugal . Steinernema carpocapsae is an entomopathogenic nematode symbiotically associated with the bacterium Xenorhabdus nematophila . This nematode secrete proteases on there secrete products. Proteases are well known catabolic functions and many tasks imposed by a parasitic life cycle. Serine proteases are the most extensively studied enzymes on host parasite interaction. Host defence mechanism including cellular and humoral reactions against invading organisms. In humoral system phenoloxidase is a key enzyme and c onsidering the parasite survival it may inhibit prophenoloxidase system, could be a key to successful parasitism. In order to study host immune suppression, the in fective third-stage (L3) S. carpocapsae nematodes were grown on 1% Galleria mellonella insect homogenate and then collect secretory products (SP). From the SP trypsin (Sc-TRYP) and chymo trypsin (Sc-CHYM) -like serine protease were identified, purified and characterized. I n vitro, 52.6 % and 63.7% prophenoloxidase suppression was observed for Sc-TRYP and Sc-CHYM respectively. Sc-TRYP could affect G. mellonella insect haemocyte causing cells to become spherical or round shape and actin filaments were detected with Phalloidintetramethylrhodamine showed some of them were disorganised in haemocytes. Sc-chym gene expression was analysed with induced and non induced insect homogenate for 6 to72 h. In vivo , purified Sc-CHYM imbibed beads showed, it could prevent haemocytes encapsulation and melanization by 12 and 24 h, respectively. The role of these proteases in host immune suppression will be discussed.

JM Barral et al. (1999) International C. elegans Meeting "UCS Domain Functions as a Myosin Assemblase"

HF Epstein, JM Barral

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International C. elegans Meeting,

1999]

Muscle thick filaments have served as models to study biological multi-protein assemblies. In C. elegans , body wall thick filaments are tubules with a core of paramyosin subfilaments coupled by filagenins, and a surface layer of two differentially localized myosin isoforms: myosin A in the center and myosin B in the polar regions (Liu et al . 1998. JCB;140:347-353). Early work suggested that the product of the unc-45 gene may function as a myosin assemblase (Epstein and Thomson. 1974. Nature;250:279-280). We have found that temperature sensitive ( ts ) mutations in the UNC-45 protein lead to a 50% decrease in the amount of thick filaments in muscle cells at the restrictive temperature. Furthermore, the filaments that assemble have abnormal structure, as evidenced by immunofluorescence and immunoelectron microscopy. Myosins A and B are scrambled throughout the filament length and upon isolation, the filaments display marked in vitro instability. Three ts mutations are amino acid substitutions in conserved residues within a predicted "UCS" domain similar to three fungal proteins: CRO1 ( P. anserina ), She4p ( S. cerevisiae ), and Rng3p ( S. pombe ) (Barral et al . 1998. JCB;143:1215-1225). These proteins were found in screens searching for mutants defective in septation ( CRO : Berteaux-Lecellier et al . 1998. EMBOJ;17:1248-1258), the segregation of molecules during budding and endocytosis ( SHE : Jansen et al . 1996. Cell;84:687-697; Wendland et al . 1996. JCB;135:1485-1500), and contractile ring formation ( RNG : Wong and Balasubramanian. 1998. MBC;9:474a). These processes all require the assembly of competent acto-myosin structures. At its amino-terminus, UNC-45 contains three TPR motifs. These are present in proteins implicated in protein folding and assembly (Frydman and Hohfeld. 1997. TIBS;22:87-92). Our current work is directed towards testing the model that the UCS domain of UNC-45 binds myosins A and B differentially, and that the TPR domain interacts with molecular chaperones, which keep myosin in an assembly-competent state until it is properly placed in the filament. UCS proteins may operate through a common mechanism, recognizing specific myosin isoforms and assembling them into structures appropriate for their particular cellular function.