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[
Bioinformation,
2007]
C. elegans C46H11.4 gene encodes a Let-23 fertility effector/regulator protein of the EGF-receptor class of the tyrosine kinase family. Alternative splicing is a major mechanism of generating protein diversity in higher eukaryotes. C. elegans genome sequencing consortium has reported three alternatively spliced transcripts of C46H11.4 gene which encodes for three hypothetical proteins namely, C46H11.4a, C46H11.4b and C46H11.4c. Using a combination of various bioinformatics tools like gene or exon finding programmes, blast searches, alignment tools etc followed by experimental validation, we report the presence of three more alternatively spliced transcripts which encode for novel hypothetical proteins C46H11.4d, C46H11.4e and C46H11.4f. These isoforms arise as a result of alternative splicing in the pre-mRNA encoded by gene C46H11.4. These novel un-reported spliced variants not only point towards the extent of alternative splicing in C. elegans genes but also hint towards the complex nature of alternative splicing.
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[
J Biol Chem,
1997]
Recent work has shown that the murine BRCA2 tumor suppressor protein interacts with the murine RAD51 protein. This interaction suggests that BRCA2 participates in DNA repair. Residues 3196-3232 of the murine BRCA2 protein were shown to be involved in this interaction. Here, we report the detailed mapping of additional domains that are involved in interactions between the human homologs of these two proteins. Through yeast two-hybrid and biochemical assays, we demonstrate that the RAD51 protein interacts specifically with the eight evolutionarily conserved BRC motifs encoded in exon 11 of brca2 and with a similar motif found in a Caenorhabditis elegans hypothetical protein. Deletion analysis demonstrates that residues 98-339 of human RAD51 interact with the 59-residue minimal region that is conserved in all BRC motifs. These data suggest that the BRC repeats function to bind RAD51.
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Verma R, Thota JR, Parmar N, Vishwakarma P, Murthy PK, Shukla PK, Kar S, Pandey S, Kushwaha V, Yadav PK, Tewari P
[
Parasitol Res,
2018]
We earlier found that F6 fraction of human filaria Brugia malayi cross-reacted with sera of Leishmania donovani infected hamsters and immunization with F6 inhibited both filarial and leishmanial infections. In the present study, we identified a 52.9-93.6 kDa fraction (Ld1) of L. donovani that cross-reacted with sera of B. malayi infected animals and investigated effect of Ld1 on filarial infection. Immunization of BALB/c mice with Ld1 facilitated B. malayi infection with remarkable increase in parasite burden. Facilitation of filarial infection was associated with downregulated cell proliferation, IL-5, IL-13, IFN-, TNF-, and IL-2 levels and upregulated IL-4 and TGF-. Ld1 exposure also suppressed MHC class-I, MHC class-II, and FcR1 expression, and phagocytosis in naive mouse macrophages, and CD4+, CD8+, and CD19+ cell population in mouse spleen. Two-dimensional electrophoresis and matrix-assisted laser desorption ionization-time of flight-mass spectrometry revealed eight proteins in Ld1: putative heat shock protein (HSP) 70-related protein 1, HSP70 mitochondrial precursor, alanine aminotransferase, 2,3-bisphosphoglycerate-independent phosphoglycerate mutase, protein disulfide isomerase, putative ATPase beta subunit, trypanothione reductase, and a hypothetical protein. HSP70 protein mitochondrial precursor and trypanothione reductase showed homology with Trypanosoma cruzi and L. donovani, respectively, and the rest 6 proteins including hypothetical protein bear homology with L. infantum. In conclusion, the present study for the first time shows that immunization with filarial cross-reactive Ld1 fraction of L. donovani facilitates filarial infection by modulating Th1 and Th2 responses. Ld1 molecules may therefore facilitate filarial infection in filaria-leishmania co-infection.
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[
Proteomics,
2004]
Proteome maps and differences of protein patterns of the synchronized larval stage L4 of the temperature-sensitive Caenorhabditis elegans (C. elegans)
glp-1 mutant (
e2144ts) were investigated after cultivation at 15degreesC (developing a normal phenotype) or 25degreesC (developing a mutated phenotype) by two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization (MALDI)-mass spectrometry. From the 183 identified protein spots six proteins were found differently expressed. The Vit-6 vitellogenin (CE28594), the hypothetical 17.2 protein (CE25224), the hypothetical 17.4 protein (CE16999), and the heat shock protein 16 kDa (CE14249) were more abundant when growing worm cultures at 25degreesC. By contrast, the nucleoside diphosphate kinase (CE09650) was found increased at 15degreesC. Most notably, the eukariotic initiation factor 5A-1 (CE00503), highly abundant at 15degreesC, was not present in cultures grown at 25degreesC. Its absence at 25degreesC can not be attributed to lack of the enzymatic machinery that is necessary for hypusinylation. Instead, a direct downstream effect of the lack of functionality of GLP-1 may cause the expression of this protein. The yolk proteins 115 kDa and 88 kDa were attributed by mass spectrometric protein structure analysis as C-terminal and N-terminal fragments of the Vit-6 vitellogin protein (CE28594), respectively. The cleavage site between both derivatives was located between R764 and A768. A conflict in the database sequences at amino acid positions 1622 and 1623 of vitellogenin-6 was solved by mass spectrometric sequence analysis. The combination of 2-DE with mass spectrometry enabled the identification of mutation-associated differences on somatic gonadal cell and germ line cell development-associated proteins.
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[
Exp Parasitol,
2019]
Adult Brugia malayi proteins with high potential as epidemiological markers, diagnostic and therapeutic targets, and/or vaccine candidates were revealed by using microfilaremic human sera and an immunoproteomic approach. They were HSP70, cytoplasmic intermediate filament protein, independent phosphoglycerate mutase, and enolase. Brugia malayi microfilaria-specific proteins that formed circulating immune complexes (ICs) were investigated. The IC-forming proteins were orthologues of hypothetical protein Bm1_12480, Pao retrotransposon peptidase family protein, uncoordinated protein 44, NAD-binding domain containing protein of the UDP-glucose/GDP-mannose dehydrogenase family which contained ankyrin repeat region, ZU5 domain with C-terminal death domain, C2 domain containing protein, and FLJ90013 protein of the eukaryotic membrane protein family. Antibodies to these proteins were not free in the microfilaremic sera, raising the possible role of the IC-forming proteins in an immune evasion mechanism of the circulating microfilariae to avoid antibody-mediated-host immunity. Moreover, detection of these ICs should be able to replace the inconvenient night blood sampling for microfilaria in an evaluation of efficacy of anti-microfilarial agents.
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[
Biochem Biophys Res Commun,
2003]
We report here the first identification and structural characterization of a eukaryotic protein with homology to the bacterial MgtE family of potential Mg(2+) transporters. This human protein, denoted solute carrier family 41 member 1 (SLC41A1), consists of 513 amino acids with an estimated molecular weight of 56kDa. Computer analysis of the protein structure reveals that the protein consists of 10 putative transmembrane domains and includes two distinct domains highly homologous to the integral membrane part of the bacterial MgtE protein family. The gene encoding SLC41A1 is found on chromosome 1 (1q31-32) and the protein coding sequence is found on 10 exons. A 5-kb long transcript is identified in various human tissues with highest expression levels in heart and testis. We have also identified 10 SLC41A1 homologs in Homo sapiens, Mus musculus, Drosophila melanogaster, Anopheles gambiae, and Caenorhabditis elegans, and propose that these hypothetical proteins belong to a novel eukaryotic gene family.
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[
International Worm Meeting,
2009]
One of the adaptive behaviors of worms in their environment is thermotaxis, by which thery migrate toward a prefered temperature. The thermotactic behavior is accomplished by choosing thermophilic or cryophilic movement depending on the surrounding temperature. However, some experimental data are inconsistent especially for thermophilic movement, which is expected to be observed in lower than prefered temperatures. There are no experimental analyses which support thermophilic movement in the individual behavior of worms. Although mathematical modeling is used to study thermotactic behavior in C. elegans, no model provides a consistent explanation for this discrepancy. Here we develop a simple mathematical model based on biased random walk, which describes population behavior. We determined the parameters of the model according to the experimental results of individual movement assays. In addition to them, we examined hypothetical bias as thermophilic drive. Our model can not regenerate some behavioral patterns of population distribution without the hypothetical bias. However our model can regenerate all the population patterns reported in past studies without any contradiction if we use the hypothetical bias into our model. In addition, our model show that thermophilic movement can be observed only when the steepness of thermal gradient is sufficiently slight. If the gradient is too steep in contrast, our model do not show a sharp contrast between the results with thermophilic bias and the results without it. Our results therefore suggest that thermophilic movement would be observed, even in individual movements, when the thermal gradient is sufficiently slight. On the contrary, thermophilic movement dissapears when the thermal gradient is too steep. The steepness of thermal gradient is thus essential to understand the past experimental studies without discrepancy.
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[
Biochim Biophys Acta,
2008]
The Dauer larva is a non-feeding alternative larval stage of some nematodes specialized for long-term survival and dispersal. In this study we compared proteome maps obtained from Dauer larvae with those from the corresponding third larval stage (L3) of the feeding life cycle of C. elegans wild-type strain N2. We demonstrate at the protein level that altered metabolism may participate in longevity determination of Dauers. We detected huge amounts of alcohol dehydrogenase (CE12212) and aldehyde dehydrogenase (CE29809) in Dauer animals, indicating highly active fermentative pathways. Inorganic pyrophosphatase (CE05448) that enables to metabolize pyrophosphate as a high-energy source was over-expressed in Dauers. An interesting differentially expressed protein was phosphatidylethanolamine-binding protein (CE38516) that was found in high abundance in samples from Dauer larvae. Protein synthesis may be lowered in Dauer animals by the reduced expression of splicing factor
rsp-3 (CE31089) and methionyl-tRNA synthase (CE34219). We observed significantly lower amounts of the pepsin-like aspartyl protease 1 (CE21681) in non-feeding Dauers, which is in agreement with reduced nutrient digestion. Finally, the hypothetical protein R08E5.2 (CE33294) was present in high abundance in L3 animals.
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Chen, Jia-Xuan, Polanowska, Jolanta, Gunsalus, Kristin, Piano, Fabio, Cipriani, Patricia, Mecenas, Desirea, Selbach, Matthias
[
International Worm Meeting,
2015]
Mapping protein-protein interactions in vivo is instrumental in deciphering the molecular mechanisms underlying animal development. We have developed a new method combining in vivo expressed GFP-tagged proteins with label-free quantitative proteomics to identify protein-protein interactions in developing C. elegans embryos. This strategy is generic and can in principle be used with any GFP-tagged protein. To test our approach we focused on eight proteins involved in essential biological processes during embryogenesis and built a pilot embryo in vivo interaction map comprising 559 interactions among 472 proteins. This network captures known biology and is highly enriched in functionally relevant interactions. We further show the utility of the map by searching for new regulators of P granule formation during embryogenesis. We discovered the worm-specific protein GEI-12 as a novel interaction partner of the DYRK kinase MBK-2 and as an important regulator of P granule dynamics and germline maintenance. This leads us to propose a hypothetical model in which the phosphorylation state of GEI-12 regulates P granule assembly and disassembly during early embryogenesis. Additionally, GEI-12 also induces granule formation in mammalian cells, suggesting that a common mechanism of ribonucleoprotein granule assembly exists in worms and humans. Our results show that in vivo interactome mapping is a powerful and versatile approach that provides unique insights into animal development.
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[
Biochem Mol Biol Int,
1993]
Oligonucleotides related to parts of a globin-like sequence in the genome of Caenorhabditis elegans were used to probe a cDNA library from the same species. A complete globin-like sequence was found in the cDNA, showing that a globin gene appears to be expressed. The hypothetical protein was compatible with the conventional globin fold but may be truncated in the B helix, as in Chironomus globin III. An intron in the codon for residue E3 in the E helix was removed in expression. An initiation codon preceded the globin but the sequence upstream (extending for 30 nucleotides to the vector ligation site) had characteristics both of the code for a protein hydrophobic leader and of a trans-spliced RNA leader. The evidence indicates that C. elegans globin has a single domain, unlike some nematodes that express two tandem globin domains in a continuous translation product, and from its sequence may be predicted to have a high affinity for oxygen.