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A Synthetic Lethal Screen Identifies a Role for Lin-44/Wnt in C. elegans Embryogenesis.

Hartin SN, Hudson ML, Yingling C, Ackley BD - PLoS ONE (2015)

Bottom Line: We found animals with LOF in both sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood.We found that the Wnt ligand, lin-44, was SynLet with sdn-1, but not ptp-3.We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, University of Kansas, Lawrence, KS, United States of America.

ABSTRACT

Background: The C. elegans proteins PTP-3/LAR-RPTP and SDN-1/Syndecan are conserved cell adhesion molecules. Loss-of-function (LOF) mutations in either ptp-3 or sdn-1 result in low penetrance embryonic developmental defects. Work from other systems has shown that syndecans can function as ligands for LAR receptors in vivo. We used double mutant analysis to test whether ptp-3 and sdn-1 function in a linear genetic pathway during C. elegans embryogenesis.

Results: We found animals with LOF in both sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood. Analysis of the survivors demonstrated that these animals had a synergistic increase in the penetrance of embryonic developmental defects. Together, these data strongly suggested PTP-3 and SDN-1 function in parallel during embryogenesis. We subsequently used RNAi to knockdown ~3,600 genes predicted to encode secreted and/or transmembrane molecules to identify genes that interacted with ptp-3 or sdn-1. We found that the Wnt ligand, lin-44, was SynLet with sdn-1, but not ptp-3. We used 4-dimensional time-lapse analysis to characterize the interaction between lin-44 and sdn-1. We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1.

Conclusions: PTP-3 and SDN-1 function in compensatory pathways during C. elegans embryonic and larval development, as simultaneous loss of both genes has dire consequences for organismal survival. The Wnt ligand lin-44 contributes to the early stages of gastrulation in parallel to sdn-1, but in a genetic pathway with ptp-3. Overall, the SynLet phenotype provides a robust platform to identify ptp-3 and sdn-1 interacting genes, as well as other genes that function in development, yet might be missed in traditional forward genetic screens.

No MeSH data available.


Related in: MedlinePlus

Epidermal junctions can be maintained in cell-adhesion mutants and RNAi treated animals.We used an AJM-1::GFP transgene (jcIs1) to examine epidermal morphology in animals being tested. A: In wild-type jcIs1 animals, AJM-1::GFP is localized to cell junctions (arrow) and outlines epidermal cells. Here a view of the dorsal epidermal cells is visible. B: A dorsal view of an sdn-1(zh20) animal showing disorganized epidermal cells in the posterior half of the embryo. C: Ventro-lateral view of a wild-type embryo just after ventral enclosure. Asterisks mark the hexagon-shaped lateral seam cells. Note the regular morphology. D: A sdn-1(zh20) animal treated with lin-44 RNAi. While some lateral seam cells (asterisks) are correctly positioned, others are grossly disorganized (note, some of these seam cell identities were made tentatively, and are based on location). Scale bar = 10 μm.
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pone.0121397.g004: Epidermal junctions can be maintained in cell-adhesion mutants and RNAi treated animals.We used an AJM-1::GFP transgene (jcIs1) to examine epidermal morphology in animals being tested. A: In wild-type jcIs1 animals, AJM-1::GFP is localized to cell junctions (arrow) and outlines epidermal cells. Here a view of the dorsal epidermal cells is visible. B: A dorsal view of an sdn-1(zh20) animal showing disorganized epidermal cells in the posterior half of the embryo. C: Ventro-lateral view of a wild-type embryo just after ventral enclosure. Asterisks mark the hexagon-shaped lateral seam cells. Note the regular morphology. D: A sdn-1(zh20) animal treated with lin-44 RNAi. While some lateral seam cells (asterisks) are correctly positioned, others are grossly disorganized (note, some of these seam cell identities were made tentatively, and are based on location). Scale bar = 10 μm.

Mentions: In wild type animals, AJM-1::GFP can be seen accumulating at cell junctions outlining the epidermal cells, starting around the lima bean stage of embryogenesis, and persisting throughout development (Fig 4). Epidermal cell junctions in wild-type animals are well organized, and only rarely display gaps or misshapen cells. In contrast, we found that both ptp-3 and sdn-1 mutants had apparent cell-shape changes consistent with defects in either cell positioning or cell polarity (Fig 4, Table 3).


A Synthetic Lethal Screen Identifies a Role for Lin-44/Wnt in C. elegans Embryogenesis.

Hartin SN, Hudson ML, Yingling C, Ackley BD - PLoS ONE (2015)

Epidermal junctions can be maintained in cell-adhesion mutants and RNAi treated animals.We used an AJM-1::GFP transgene (jcIs1) to examine epidermal morphology in animals being tested. A: In wild-type jcIs1 animals, AJM-1::GFP is localized to cell junctions (arrow) and outlines epidermal cells. Here a view of the dorsal epidermal cells is visible. B: A dorsal view of an sdn-1(zh20) animal showing disorganized epidermal cells in the posterior half of the embryo. C: Ventro-lateral view of a wild-type embryo just after ventral enclosure. Asterisks mark the hexagon-shaped lateral seam cells. Note the regular morphology. D: A sdn-1(zh20) animal treated with lin-44 RNAi. While some lateral seam cells (asterisks) are correctly positioned, others are grossly disorganized (note, some of these seam cell identities were made tentatively, and are based on location). Scale bar = 10 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4418752&req=5

pone.0121397.g004: Epidermal junctions can be maintained in cell-adhesion mutants and RNAi treated animals.We used an AJM-1::GFP transgene (jcIs1) to examine epidermal morphology in animals being tested. A: In wild-type jcIs1 animals, AJM-1::GFP is localized to cell junctions (arrow) and outlines epidermal cells. Here a view of the dorsal epidermal cells is visible. B: A dorsal view of an sdn-1(zh20) animal showing disorganized epidermal cells in the posterior half of the embryo. C: Ventro-lateral view of a wild-type embryo just after ventral enclosure. Asterisks mark the hexagon-shaped lateral seam cells. Note the regular morphology. D: A sdn-1(zh20) animal treated with lin-44 RNAi. While some lateral seam cells (asterisks) are correctly positioned, others are grossly disorganized (note, some of these seam cell identities were made tentatively, and are based on location). Scale bar = 10 μm.
Mentions: In wild type animals, AJM-1::GFP can be seen accumulating at cell junctions outlining the epidermal cells, starting around the lima bean stage of embryogenesis, and persisting throughout development (Fig 4). Epidermal cell junctions in wild-type animals are well organized, and only rarely display gaps or misshapen cells. In contrast, we found that both ptp-3 and sdn-1 mutants had apparent cell-shape changes consistent with defects in either cell positioning or cell polarity (Fig 4, Table 3).

Bottom Line: We found animals with LOF in both sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood.We found that the Wnt ligand, lin-44, was SynLet with sdn-1, but not ptp-3.We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, University of Kansas, Lawrence, KS, United States of America.

ABSTRACT

Background: The C. elegans proteins PTP-3/LAR-RPTP and SDN-1/Syndecan are conserved cell adhesion molecules. Loss-of-function (LOF) mutations in either ptp-3 or sdn-1 result in low penetrance embryonic developmental defects. Work from other systems has shown that syndecans can function as ligands for LAR receptors in vivo. We used double mutant analysis to test whether ptp-3 and sdn-1 function in a linear genetic pathway during C. elegans embryogenesis.

Results: We found animals with LOF in both sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood. Analysis of the survivors demonstrated that these animals had a synergistic increase in the penetrance of embryonic developmental defects. Together, these data strongly suggested PTP-3 and SDN-1 function in parallel during embryogenesis. We subsequently used RNAi to knockdown ~3,600 genes predicted to encode secreted and/or transmembrane molecules to identify genes that interacted with ptp-3 or sdn-1. We found that the Wnt ligand, lin-44, was SynLet with sdn-1, but not ptp-3. We used 4-dimensional time-lapse analysis to characterize the interaction between lin-44 and sdn-1. We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1.

Conclusions: PTP-3 and SDN-1 function in compensatory pathways during C. elegans embryonic and larval development, as simultaneous loss of both genes has dire consequences for organismal survival. The Wnt ligand lin-44 contributes to the early stages of gastrulation in parallel to sdn-1, but in a genetic pathway with ptp-3. Overall, the SynLet phenotype provides a robust platform to identify ptp-3 and sdn-1 interacting genes, as well as other genes that function in development, yet might be missed in traditional forward genetic screens.

No MeSH data available.


Related in: MedlinePlus