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Doctoral Thesis
DOI
https://doi.org/10.11606/T.42.2009.tde-03112009-104538
Document
Author
Full name
Daniela Peres Almenara
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2009
Supervisor
Committee
Winter, Carlos Eduardo (President)
Boscardin, Silvia Beatriz
Gorab, Eduardo
Menck, Carlos Frederico Martins
Monesi, Nadia
Title in Portuguese
Estudo das vitelinas VT1 e YP170B dos nematoides rabditídeos Oscheius tipulae e Caenorhabditis elegans: aspectos estruturais e funcionais.
Keywords in Portuguese
Caenorhabditis elegans
Oscheius tipulae
Ligand-blotting
MTP
Nematóides (Estrutura)
Parasitologia
RNAi
Vitelogenina
Abstract in Portuguese
A região N-terminal de OTI-VIT-1 foi expressa e os polipeptídeos recombinantes foram purificados. OTI-VIT-1 pode ser homólogo da vitelina YP170B de C. elegans. Foram identificados um intron na região 5´ e dois na região 3´ do gene Oti-vit-1. Antissoro monoespecífico para PVIT1HisC confirmou que o gene Oti-vit-1 codifica VT1. O polipeptídeo recombinante P40-H, correspondente à região N-terminal da proteína OTI-VIT-6 interage com um polipeptídeo de aproximadamente 100 kDa (P100) presente em extratos proteicos totais de O. tipulae. Estudamos também o papel da Proteína Microssômica Transportadora de Triglicerídeos (MTP) na biossíntese de Vitelogenina do nematoide C. elegans. Ensaios de RNAi em C. elegans, utilizando parte da sequência do gene da MTP (Cel-dsc-4) foram realizados nas linhagens N2 e DH1033. Microscopia de fluorescência de vermes adultos da linhagem DH1033, submetidos a RNAi, mostrou acúmulo de YP170B::GFP no interior dos enterócitos. Este acúmulo sugere a participação da MTP na secreção de VTG. Análise imunológica da vitelogenina nestes mesmos vermes não detectaram alterações no processamento de CEL-VIT-6, sugerindo que o mesmo ocorra não só no pseudoceloma, mas também no interior dos enterócitos.
Title in English
Structural and functional analysis of VT1 and YP170B vitellins from the Rhabditid nematodes Oscheius tipulae and Caenorhabditis elegans.
Keywords in English
Caenorhabditis elegans
Oscheius tipulae
Ligand-blotting
MTP
Nematodes (Structure)
Parasitology
RNAi
Vitellogenin
Abstract in English
The N-terminal region of OTI-VIT-1 was expressed and the recombinant polypeptides were purified. OTI-VIT-1 may be homologous to the vitellin YP170B from C. elegans. We identified an intron in the 5 'region and two in 3' region from Oti-vit-1. Monospecific antisera to PVIT1HisC confirmed that the gene Oti-vit-1 encodes VT1. The recombinant polypeptide P40-H, corresponding to the N-terminal region of the protein OTI-VIT-6, interacts with a polypeptide of approximately 100 kDa (P100) present in total protein extracts of O. tipulae. The role of microsomal triglyceride transfer protein (MTP) in the biosynthesis of vitellogenin was studied in the nematode C. elegans. Trials of RNAi in C. elegans, using the sequence of the MTP gene (Cel-dsc-4) were performed in the strains N2 and DH1033. Fluorescence microscopy of adult worms of strain DH1033, subjected to RNAi, showed accumulation of YP170B:: GFP within the enterocytes. This accumulation suggests the involvement of MTP in the secretion of VTG. Analysis using anti-vitellogenin immune serum did not detect changes in the processing of CEL-VIT-6, suggesting that it occurs not only in pseudocoelom but also within the enterocytes.
 
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Publishing Date
2010-02-12
 
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