Limits...
Caenorhabditis elegans is a useful model for anthelmintic discovery.

Burns AR, Luciani GM, Musso G, Bagg R, Yeo M, Zhang Y, Rajendran L, Glavin J, Hunter R, Redman E, Stasiuk S, Schertzberg M, Angus McQuibban G, Caffrey CR, Cutler SR, Tyers M, Giaever G, Nislow C, Fraser AG, MacRae CA, Gilleard J, Roy PJ - Nat Commun (2015)

Bottom Line: We then rescreen our hits in two parasitic nematode species and two vertebrate models (HEK293 cells and zebrafish), and identify 30 structurally distinct anthelmintic lead molecules.We identify the target of one lead with nematode specificity and nanomolar potency as complex II of the electron transport chain.This work establishes C. elegans as an effective and cost-efficient model system for anthelmintic discovery.

View Article: PubMed Central - PubMed

Affiliation: The Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, Ontario, Canada M5S 3E1.

ABSTRACT
Parasitic nematodes infect one quarter of the world's population and impact all humans through widespread infection of crops and livestock. Resistance to current anthelmintics has prompted the search for new drugs. Traditional screens that rely on parasitic worms are costly and labour intensive and target-based approaches have failed to yield novel anthelmintics. Here, we present our screen of 67,012 compounds to identify those that kill the non-parasitic nematode Caenorhabditis elegans. We then rescreen our hits in two parasitic nematode species and two vertebrate models (HEK293 cells and zebrafish), and identify 30 structurally distinct anthelmintic lead molecules. Genetic screens of 19 million C. elegans mutants reveal those nematicides for which the generation of resistance is and is not likely. We identify the target of one lead with nematode specificity and nanomolar potency as complex II of the electron transport chain. This work establishes C. elegans as an effective and cost-efficient model system for anthelmintic discovery.

No MeSH data available.


Related in: MedlinePlus

Complex II residues that are mutated in the wact-11 family resistant mutants cluster near the ubiquinone-binding site (Q-site).(a) Rendering of the crystal structure of Ascaris suum Complex II bound to the Q-site inhibitor flutolanil (PDB: 3VRB). The side chains of the 14 orthologous residues that are mutated in the wact-11-family resistant mutants are shown as opaque spheres. The atoms of the bound flutolanil molecule are shown as orange-coloured opaque spheres. (b) Close-up view of flutolanil bound at the Q-site of Complex II from Ascaris suum. The 12 residues shown are no more than 4 Å away from flutolanil, and make up the flutolanil binding pocket. Intermolecular distances are indicated with bidirectional arrows. The dashed line represents a hydrogen bond (H-bond) interaction. Only those H-bonds that occur between Complex II residues and flutolanil are shown; H-bonds that occur between residues of Complex II were omitted for clarity. Bound cofactors, and a bound fumarate molecule, were also omitted for clarity.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4491176&req=5

f4: Complex II residues that are mutated in the wact-11 family resistant mutants cluster near the ubiquinone-binding site (Q-site).(a) Rendering of the crystal structure of Ascaris suum Complex II bound to the Q-site inhibitor flutolanil (PDB: 3VRB). The side chains of the 14 orthologous residues that are mutated in the wact-11-family resistant mutants are shown as opaque spheres. The atoms of the bound flutolanil molecule are shown as orange-coloured opaque spheres. (b) Close-up view of flutolanil bound at the Q-site of Complex II from Ascaris suum. The 12 residues shown are no more than 4 Å away from flutolanil, and make up the flutolanil binding pocket. Intermolecular distances are indicated with bidirectional arrows. The dashed line represents a hydrogen bond (H-bond) interaction. Only those H-bonds that occur between Complex II residues and flutolanil are shown; H-bonds that occur between residues of Complex II were omitted for clarity. Bound cofactors, and a bound fumarate molecule, were also omitted for clarity.

Mentions: A number of Q-site inhibitors are used as fungicides and interest in their use against nematodes is growing272829. For example, flutolanil has been shown to inhibit complex II from the parasitic nematode Ascaris suum in vitro, and a co-crystal structure of flutolanil with this complex has been solved3031. We rendered an image of this crystal structure and highlighted the corresponding 14 orthologous residues that are mutated in the wact-11-resistant C. elegans mutants (Fig. 4a). Despite residing in three distinct proteins, all the 14 residues cluster around the Q-site where flutolanil is bound. Furthermore, of the 12 residues that are within 4 angstroms of flutolanil's central mass, four are mutated in our wact-11-family resistant mutants (Fig. 4b). Finally, the most frequently mutated residue in our screen is in SDHC-1's R74, which corresponds to Ascaris' R89 of SDHC that likely makes electrostatic contacts with the benzene ring of flutolanil's 2-trifluoromethyl-benzamide group3031. Like flutolanil, wact-11 also has a 2-trifluoromethyl-benzamide group, and all wact-11 family members have the benzamide moiety. Taken together, our observations suggest that the wact-11-family kills nematodes by binding the Q-site of complex II, and consequently disrupts the interaction of ubiquinone with the complex. Furthermore, the viability of the resistant mutants suggests that the missense mutations alter the Q-site in a way that preserves its function.


Caenorhabditis elegans is a useful model for anthelmintic discovery.

Burns AR, Luciani GM, Musso G, Bagg R, Yeo M, Zhang Y, Rajendran L, Glavin J, Hunter R, Redman E, Stasiuk S, Schertzberg M, Angus McQuibban G, Caffrey CR, Cutler SR, Tyers M, Giaever G, Nislow C, Fraser AG, MacRae CA, Gilleard J, Roy PJ - Nat Commun (2015)

Complex II residues that are mutated in the wact-11 family resistant mutants cluster near the ubiquinone-binding site (Q-site).(a) Rendering of the crystal structure of Ascaris suum Complex II bound to the Q-site inhibitor flutolanil (PDB: 3VRB). The side chains of the 14 orthologous residues that are mutated in the wact-11-family resistant mutants are shown as opaque spheres. The atoms of the bound flutolanil molecule are shown as orange-coloured opaque spheres. (b) Close-up view of flutolanil bound at the Q-site of Complex II from Ascaris suum. The 12 residues shown are no more than 4 Å away from flutolanil, and make up the flutolanil binding pocket. Intermolecular distances are indicated with bidirectional arrows. The dashed line represents a hydrogen bond (H-bond) interaction. Only those H-bonds that occur between Complex II residues and flutolanil are shown; H-bonds that occur between residues of Complex II were omitted for clarity. Bound cofactors, and a bound fumarate molecule, were also omitted for clarity.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4491176&req=5

f4: Complex II residues that are mutated in the wact-11 family resistant mutants cluster near the ubiquinone-binding site (Q-site).(a) Rendering of the crystal structure of Ascaris suum Complex II bound to the Q-site inhibitor flutolanil (PDB: 3VRB). The side chains of the 14 orthologous residues that are mutated in the wact-11-family resistant mutants are shown as opaque spheres. The atoms of the bound flutolanil molecule are shown as orange-coloured opaque spheres. (b) Close-up view of flutolanil bound at the Q-site of Complex II from Ascaris suum. The 12 residues shown are no more than 4 Å away from flutolanil, and make up the flutolanil binding pocket. Intermolecular distances are indicated with bidirectional arrows. The dashed line represents a hydrogen bond (H-bond) interaction. Only those H-bonds that occur between Complex II residues and flutolanil are shown; H-bonds that occur between residues of Complex II were omitted for clarity. Bound cofactors, and a bound fumarate molecule, were also omitted for clarity.
Mentions: A number of Q-site inhibitors are used as fungicides and interest in their use against nematodes is growing272829. For example, flutolanil has been shown to inhibit complex II from the parasitic nematode Ascaris suum in vitro, and a co-crystal structure of flutolanil with this complex has been solved3031. We rendered an image of this crystal structure and highlighted the corresponding 14 orthologous residues that are mutated in the wact-11-resistant C. elegans mutants (Fig. 4a). Despite residing in three distinct proteins, all the 14 residues cluster around the Q-site where flutolanil is bound. Furthermore, of the 12 residues that are within 4 angstroms of flutolanil's central mass, four are mutated in our wact-11-family resistant mutants (Fig. 4b). Finally, the most frequently mutated residue in our screen is in SDHC-1's R74, which corresponds to Ascaris' R89 of SDHC that likely makes electrostatic contacts with the benzene ring of flutolanil's 2-trifluoromethyl-benzamide group3031. Like flutolanil, wact-11 also has a 2-trifluoromethyl-benzamide group, and all wact-11 family members have the benzamide moiety. Taken together, our observations suggest that the wact-11-family kills nematodes by binding the Q-site of complex II, and consequently disrupts the interaction of ubiquinone with the complex. Furthermore, the viability of the resistant mutants suggests that the missense mutations alter the Q-site in a way that preserves its function.

Bottom Line: We then rescreen our hits in two parasitic nematode species and two vertebrate models (HEK293 cells and zebrafish), and identify 30 structurally distinct anthelmintic lead molecules.We identify the target of one lead with nematode specificity and nanomolar potency as complex II of the electron transport chain.This work establishes C. elegans as an effective and cost-efficient model system for anthelmintic discovery.

View Article: PubMed Central - PubMed

Affiliation: The Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, Ontario, Canada M5S 3E1.

ABSTRACT
Parasitic nematodes infect one quarter of the world's population and impact all humans through widespread infection of crops and livestock. Resistance to current anthelmintics has prompted the search for new drugs. Traditional screens that rely on parasitic worms are costly and labour intensive and target-based approaches have failed to yield novel anthelmintics. Here, we present our screen of 67,012 compounds to identify those that kill the non-parasitic nematode Caenorhabditis elegans. We then rescreen our hits in two parasitic nematode species and two vertebrate models (HEK293 cells and zebrafish), and identify 30 structurally distinct anthelmintic lead molecules. Genetic screens of 19 million C. elegans mutants reveal those nematicides for which the generation of resistance is and is not likely. We identify the target of one lead with nematode specificity and nanomolar potency as complex II of the electron transport chain. This work establishes C. elegans as an effective and cost-efficient model system for anthelmintic discovery.

No MeSH data available.


Related in: MedlinePlus