Characterization of class II fumarase from Schistosoma mansoni provides the molecular basis for selective inhibition

Cardoso, Iara Aimê; Souza, Ariani Impieri de; Burgess, Adam Muslem George; Chalmers, Iain W.; Hoffmann, Karl F.; Nonato, Maria Cristina

doi:10.1016/j.ijbiomac.2021.01.180

Cited by 3 publications

(3 citation statements)

References 74 publications

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“…To determine smp_089290 knockdown (KD) in adult worms following RNAi and CRISPR/Cas9 programmed knockout, smp_089290 transcript levels in siRNA treated samples (at 48 hr post electroporation) and CRISPR/Cas9 plasmid treated samples (at seven days post electroporation) were quantified relative to smat1 as described previously [ 64 ]. The Geometric mean method was used to quantify smp_089290 transcript levels in freshly perfused male and female schistosomes as described previously [ 65 , 66 ]. Melting curves (dissociation curves) were generated for each qRT-PCR analysis to verify the amplification of one product only.…”

Section: Methodsmentioning

confidence: 99%

Schistosoma mansoni α-N-acetylgalactosaminidase (SmNAGAL) regulates coordinated parasite movement and egg production

et al. 2022

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α-galactosidase (α-GAL) and α-N-acetylgalactosaminidase (α-NAGAL) are two glycosyl hydrolases responsible for maintaining cellular homeostasis by regulating glycan substrates on proteins and lipids. Mutations in the human genes encoding either enzyme lead to neurological and neuromuscular impairments seen in both Fabry- and Schindler/Kanzaki- diseases. Here, we investigate whether the parasitic blood fluke Schistosoma mansoni, responsible for the neglected tropical disease schistosomiasis, also contains functionally important α-GAL and α-NAGAL proteins. As infection, parasite maturation and host interactions are all governed by carefully-regulated glycosylation processes, inhibiting S. mansoni’s α-GAL and α-NAGAL activities could lead to the development of novel chemotherapeutics. Sequence and phylogenetic analyses of putative α-GAL/α-NAGAL protein types showed Smp_089290 to be the only S. mansoni protein to contain the functional amino acid residues necessary for α-GAL/α-NAGAL substrate cleavage. Both α-GAL and α-NAGAL enzymatic activities were higher in females compared to males (p<0.05; α-NAGAL > α-GAL), which was consistent with smp_089290’s female biased expression. Spatial localisation of smp_089290 revealed accumulation in parenchymal cells, neuronal cells, and the vitellaria and mature vitellocytes of the adult schistosome. siRNA-mediated knockdown (>90%) of smp_089290 in adult worms significantly inhibited α-NAGAL activity when compared to control worms (siLuc treated males, p<0.01; siLuc treated females, p<0.05). No significant reductions in α-GAL activities were observed in the same extracts. Despite this, decreases in α-NAGAL activities correlated with a significant inhibition in adult worm motility as well as in egg production. Programmed CRISPR/Cas9 editing of smp_089290 in adult worms confirmed the egg reduction phenotype. Based on these results, Smp_089290 was determined to act predominantly as an α-NAGAL (hereafter termed SmNAGAL) in schistosome parasites where it participates in coordinating movement and oviposition processes. Further characterisation of SmNAGAL and other functionally important glycosyl hydrolases may lead to the development of a novel anthelmintic class of compounds.

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Section: Methodsmentioning

confidence: 99%

Schistosoma mansoni α-N-acetylgalactosaminidase (SmNAGAL) regulates coordinated parasite movement and egg production

et al. 2022

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“…Schistosomiasis is an important but often neglected tropical disease that poses a serious threat to the health of more than 250 million people worldwide [ 1 , 2 , 3 ]. Schistosomiasis is mainly caused by Schistosoma hematobium , S. mansoni , and S. japonicum [ 4 , 5 ].…”

Section: Introductionmentioning

confidence: 99%

Knocking Down Gm16685 Decreases Liver Granuloma in Murine Schistosomiasis Japonica

et al. 2023

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Long noncoding RNAs (lncRNAs) can regulate key genes and pathways in liver disease development. Moreover, macrophages are speculated to play an important role in regulating granulomatous inflammation during schistosomiasis. However, the role of lncRNAs in the formation of liver granulomas by influencing the polarization of macrophages in Schistosoma japonicum infection is unclear. Our study aimed to determine whether lncRNAs can play a role in S. japonicum-induced hepatic egg granulomas and elucidate their effect on macrophages. We established S. japonicum infection models and screened the target lncRNA Gm16685 highly expressed in schistosomiasis mice using high-throughput sequencing. Hematoxylin and eosin staining revealed that the knockdown of Gm16685 reduced the area of egg granulomas. Moreover, M1 macrophage factor genes were significantly downregulated in Gm16685 knockdown livers. Meanwhile, M2 macrophage factor genes were significantly upregulated, which was consistent with the protein detection results. Hepatocytes, hepatic stellate cells, and macrophages were isolated from mouse models infected with S. japonicum, with Gm16685 being significantly upregulated in macrophages. Moreover, the knockdown of Gm16685 in RAW264.7 cells revealed similar results to in liver tissue. RNA fluorescence in situ hybridization (FISH) and nucleocytoplasmic separation experiments revealed that Gm16685 was predominantly localized in the cytoplasm of cells. We found that miR-205-5p was upregulated after Gm16685 was knocked down. After overexpression of miR-205-5p, the expression of Gm16685 and inflammatory factors was significantly downregulated. These results indicate that Gm16685 can participate in the pathogenesis of hepatic disease in schistosomiasis and promote M1 macrophage polarization by regulating miR-205-5p. Thus, our study may provide a new target for schistosomiasis japonica treatment.

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“…The second pillar of CRAFT is the target library. Here, we have cultivated expertise in heterologous production and characterization of target proteins, − including the international recognition for our extensive work on the drug target dihydroorotate dehydrogenase . This encompasses not only biochemical, biophysical, and structural characterization, which provide essential insights for mapping function, but also the development of suitable fragment screening protocols.…”

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confidence: 99%

Introducing CRAFT: The Center for Research and Advancement in Fragments and molecular Targets

Andrade,

Nonato,

da Silva Emery

2024

ACS Med. Chem. Lett.

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We introduce the Center for Research and Advancement in Fragments and Molecular Targets (CRAFT), a pioneering research center established in 2021 through a collaboration between the University of Saõ Paulo (USP) and the Federal University of Goiaś (UFG). CRAFT integrates fragment-based drug discovery (FBDD), artificial intelligence (AI), and structural biology to develop novel therapeutic strategies. We have created fragment and target libraries and utilize AI models to streamline the drug discovery process. We invite the global scientific community to collaborate with us in addressing neglected diseases, with the goal of enhancing research capabilities and fostering scientific innovation across Latin America.

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Characterization of class II fumarase from Schistosoma mansoni provides the molecular basis for selective inhibition

Cited by 3 publications

References 74 publications

Schistosoma mansoni α-N-acetylgalactosaminidase (SmNAGAL) regulates coordinated parasite movement and egg production

Schistosoma mansoni α-N-acetylgalactosaminidase (SmNAGAL) regulates coordinated parasite movement and egg production

Knocking Down Gm16685 Decreases Liver Granuloma in Murine Schistosomiasis Japonica

Introducing CRAFT: The Center for Research and Advancement in Fragments and molecular Targets

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