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Glypican Is a Modulator of Netrin-Mediated Axon Guidance.

Blanchette CR, Perrat PN, Thackeray A, Bénard CY - PLoS Biol. (2015)

Bottom Line: However, the mechanisms regulating netrin and its receptors in the extracellular milieu are largely unknown.Here we demonstrate that in Caenorhabditis elegans, LON-2/glypican, a heparan sulfate proteoglycan, modulates UNC-6/netrin signaling and may do this through interactions with the UNC-40/DCC receptor.We also find that LON-2/glypican functions from the epidermal substrate cells to guide axons, and we provide evidence that LON-2/glypican associates with UNC-40/DCC receptor-expressing cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

ABSTRACT
Netrin is a key axon guidance cue that orients axon growth during neural circuit formation. However, the mechanisms regulating netrin and its receptors in the extracellular milieu are largely unknown. Here we demonstrate that in Caenorhabditis elegans, LON-2/glypican, a heparan sulfate proteoglycan, modulates UNC-6/netrin signaling and may do this through interactions with the UNC-40/DCC receptor. We show that developing axons misorient in the absence of LON-2/glypican when the SLT-1/slit guidance pathway is compromised and that LON-2/glypican functions in both the attractive and repulsive UNC-6/netrin pathways. We find that the core LON-2/glypican protein, lacking its heparan sulfate chains, and secreted forms of LON-2/glypican are functional in axon guidance. We also find that LON-2/glypican functions from the epidermal substrate cells to guide axons, and we provide evidence that LON-2/glypican associates with UNC-40/DCC receptor-expressing cells. We propose that LON-2/glypican acts as a modulator of UNC-40/DCC-mediated guidance to fine-tune axonal responses to UNC-6/netrin signals during migration.

No MeSH data available.


Related in: MedlinePlus

lon-2/glypican functions in the repulsive unc-6/netrin guidance pathway.(A) Schematics of the migration path of the DTCs in the wild type and examples of defective DTC migration in lon-2/glypican mutants (the anterior and the posterior DTCs exhibit similar defects). In wild-type animals, the DTCs migrate away from the vulva along the anteroposterior axis [1], then turn dorsally [2], and turn again to migrate towards the midbody region [3]. Loss of lon-2/glypican leads to defective DTC guidance, including a failure to migrate dorsally, premature dorsal turning, or a failure to remain dorsal. (B) Quantification of the DTC migration defects in lon-2/glypican mutants and rescue by lon-2(+) (see S7 Table). For each transgenic line, transgenic animals were compared to nontransgenic sibling controls. Complete loss of lon-2/glypican does not enhance the defects of the unc-6/netrin  mutants or those of the  mutants for unc-5/UNC5 and unc-40/DCC, suggesting that lon-2/glypican functions in the same guidance pathway as unc-5/UNC5, unc-40/DCC, and unc-6/netrin (see S6 Table). (C) The axons of the GABAergic motorneurons project dorsally from the ventral midline towards the dorsal nerve cord. unc-6/netrin, unc-5/UNC5, and unc-40/DCC are required for this dorsal guidance of GABAergic axons. Complete loss of lon-2/glypican does not enhance the partially penetrant defects of unc-40/DCC  mutants, suggesting that lon-2/glypican functions in the same pathway as unc-40/DCC and unc-6/netrin to guide axons dorsally (see S5 Table). Error bars are standard error of the proportion. Asterisks denote significant difference: *** p ≤ 0.001 and * p ≤ 0.05 (z-tests, p-values were corrected by multiplying by the number of comparisons). ns, not significant.
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pbio.1002183.g002: lon-2/glypican functions in the repulsive unc-6/netrin guidance pathway.(A) Schematics of the migration path of the DTCs in the wild type and examples of defective DTC migration in lon-2/glypican mutants (the anterior and the posterior DTCs exhibit similar defects). In wild-type animals, the DTCs migrate away from the vulva along the anteroposterior axis [1], then turn dorsally [2], and turn again to migrate towards the midbody region [3]. Loss of lon-2/glypican leads to defective DTC guidance, including a failure to migrate dorsally, premature dorsal turning, or a failure to remain dorsal. (B) Quantification of the DTC migration defects in lon-2/glypican mutants and rescue by lon-2(+) (see S7 Table). For each transgenic line, transgenic animals were compared to nontransgenic sibling controls. Complete loss of lon-2/glypican does not enhance the defects of the unc-6/netrin mutants or those of the mutants for unc-5/UNC5 and unc-40/DCC, suggesting that lon-2/glypican functions in the same guidance pathway as unc-5/UNC5, unc-40/DCC, and unc-6/netrin (see S6 Table). (C) The axons of the GABAergic motorneurons project dorsally from the ventral midline towards the dorsal nerve cord. unc-6/netrin, unc-5/UNC5, and unc-40/DCC are required for this dorsal guidance of GABAergic axons. Complete loss of lon-2/glypican does not enhance the partially penetrant defects of unc-40/DCC mutants, suggesting that lon-2/glypican functions in the same pathway as unc-40/DCC and unc-6/netrin to guide axons dorsally (see S5 Table). Error bars are standard error of the proportion. Asterisks denote significant difference: *** p ≤ 0.001 and * p ≤ 0.05 (z-tests, p-values were corrected by multiplying by the number of comparisons). ns, not significant.

Mentions: In addition to unc-6/netrin acting as an attractive cue for cells expressing the unc-40/DCC receptor in ventral guidance, unc-6/netrin also acts as a repulsive cue for cells expressing both the unc-5/UNC5 and unc-40/DCC receptors, which together mediate dorsal guidance away from unc-6/netrin [4–6]. To address whether lon-2/glypican functions in unc-6/netrin-mediated repulsive guidance as well, we examined the dorsal migration of the distal tip cells (DTCs) and of the GABAergic motorneuron axons [4,11]. We found that lon-2/glypican single mutants are defective in dorsal DTC migrations (Fig 2A and 2B) and that the complete loss of lon-2/glypican did not enhance the dorsal DTC migration defects of unc-6/netrin, unc-40/DCC, or unc-5/UNC5 mutants (Fig 2B), indicating that lon-2/glypican functions in the unc-6/netrin-repulsive guidance pathway as well. Similarly, complete loss of lon-2/glypican did not enhance the defects of unc-40/DCC mutants in the dorsal guidance of motorneuron axons (Fig 2C). Given that loss of lon-2/glypican enhances the motorneuron axon guidance defects of sdn-1 mutants as shown in [17], lon-2/glypican plays a role in the dorsal guidance of motorneuron axons. The lack of enhancement of the defects in the dorsal guidance of motorneuron axons of unc-40/DCC mutants by loss of lon-2/glypican further supports that lon-2/glypican functions in the unc-6/netrin pathway mediating dorsal guidance. Thus, lon-2/glypican may modulate unc-6/netrin signaling not only during attractive guidance but also during repulsive guidance.


Glypican Is a Modulator of Netrin-Mediated Axon Guidance.

Blanchette CR, Perrat PN, Thackeray A, Bénard CY - PLoS Biol. (2015)

lon-2/glypican functions in the repulsive unc-6/netrin guidance pathway.(A) Schematics of the migration path of the DTCs in the wild type and examples of defective DTC migration in lon-2/glypican mutants (the anterior and the posterior DTCs exhibit similar defects). In wild-type animals, the DTCs migrate away from the vulva along the anteroposterior axis [1], then turn dorsally [2], and turn again to migrate towards the midbody region [3]. Loss of lon-2/glypican leads to defective DTC guidance, including a failure to migrate dorsally, premature dorsal turning, or a failure to remain dorsal. (B) Quantification of the DTC migration defects in lon-2/glypican mutants and rescue by lon-2(+) (see S7 Table). For each transgenic line, transgenic animals were compared to nontransgenic sibling controls. Complete loss of lon-2/glypican does not enhance the defects of the unc-6/netrin  mutants or those of the  mutants for unc-5/UNC5 and unc-40/DCC, suggesting that lon-2/glypican functions in the same guidance pathway as unc-5/UNC5, unc-40/DCC, and unc-6/netrin (see S6 Table). (C) The axons of the GABAergic motorneurons project dorsally from the ventral midline towards the dorsal nerve cord. unc-6/netrin, unc-5/UNC5, and unc-40/DCC are required for this dorsal guidance of GABAergic axons. Complete loss of lon-2/glypican does not enhance the partially penetrant defects of unc-40/DCC  mutants, suggesting that lon-2/glypican functions in the same pathway as unc-40/DCC and unc-6/netrin to guide axons dorsally (see S5 Table). Error bars are standard error of the proportion. Asterisks denote significant difference: *** p ≤ 0.001 and * p ≤ 0.05 (z-tests, p-values were corrected by multiplying by the number of comparisons). ns, not significant.
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pbio.1002183.g002: lon-2/glypican functions in the repulsive unc-6/netrin guidance pathway.(A) Schematics of the migration path of the DTCs in the wild type and examples of defective DTC migration in lon-2/glypican mutants (the anterior and the posterior DTCs exhibit similar defects). In wild-type animals, the DTCs migrate away from the vulva along the anteroposterior axis [1], then turn dorsally [2], and turn again to migrate towards the midbody region [3]. Loss of lon-2/glypican leads to defective DTC guidance, including a failure to migrate dorsally, premature dorsal turning, or a failure to remain dorsal. (B) Quantification of the DTC migration defects in lon-2/glypican mutants and rescue by lon-2(+) (see S7 Table). For each transgenic line, transgenic animals were compared to nontransgenic sibling controls. Complete loss of lon-2/glypican does not enhance the defects of the unc-6/netrin mutants or those of the mutants for unc-5/UNC5 and unc-40/DCC, suggesting that lon-2/glypican functions in the same guidance pathway as unc-5/UNC5, unc-40/DCC, and unc-6/netrin (see S6 Table). (C) The axons of the GABAergic motorneurons project dorsally from the ventral midline towards the dorsal nerve cord. unc-6/netrin, unc-5/UNC5, and unc-40/DCC are required for this dorsal guidance of GABAergic axons. Complete loss of lon-2/glypican does not enhance the partially penetrant defects of unc-40/DCC mutants, suggesting that lon-2/glypican functions in the same pathway as unc-40/DCC and unc-6/netrin to guide axons dorsally (see S5 Table). Error bars are standard error of the proportion. Asterisks denote significant difference: *** p ≤ 0.001 and * p ≤ 0.05 (z-tests, p-values were corrected by multiplying by the number of comparisons). ns, not significant.
Mentions: In addition to unc-6/netrin acting as an attractive cue for cells expressing the unc-40/DCC receptor in ventral guidance, unc-6/netrin also acts as a repulsive cue for cells expressing both the unc-5/UNC5 and unc-40/DCC receptors, which together mediate dorsal guidance away from unc-6/netrin [4–6]. To address whether lon-2/glypican functions in unc-6/netrin-mediated repulsive guidance as well, we examined the dorsal migration of the distal tip cells (DTCs) and of the GABAergic motorneuron axons [4,11]. We found that lon-2/glypican single mutants are defective in dorsal DTC migrations (Fig 2A and 2B) and that the complete loss of lon-2/glypican did not enhance the dorsal DTC migration defects of unc-6/netrin, unc-40/DCC, or unc-5/UNC5 mutants (Fig 2B), indicating that lon-2/glypican functions in the unc-6/netrin-repulsive guidance pathway as well. Similarly, complete loss of lon-2/glypican did not enhance the defects of unc-40/DCC mutants in the dorsal guidance of motorneuron axons (Fig 2C). Given that loss of lon-2/glypican enhances the motorneuron axon guidance defects of sdn-1 mutants as shown in [17], lon-2/glypican plays a role in the dorsal guidance of motorneuron axons. The lack of enhancement of the defects in the dorsal guidance of motorneuron axons of unc-40/DCC mutants by loss of lon-2/glypican further supports that lon-2/glypican functions in the unc-6/netrin pathway mediating dorsal guidance. Thus, lon-2/glypican may modulate unc-6/netrin signaling not only during attractive guidance but also during repulsive guidance.

Bottom Line: However, the mechanisms regulating netrin and its receptors in the extracellular milieu are largely unknown.Here we demonstrate that in Caenorhabditis elegans, LON-2/glypican, a heparan sulfate proteoglycan, modulates UNC-6/netrin signaling and may do this through interactions with the UNC-40/DCC receptor.We also find that LON-2/glypican functions from the epidermal substrate cells to guide axons, and we provide evidence that LON-2/glypican associates with UNC-40/DCC receptor-expressing cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

ABSTRACT
Netrin is a key axon guidance cue that orients axon growth during neural circuit formation. However, the mechanisms regulating netrin and its receptors in the extracellular milieu are largely unknown. Here we demonstrate that in Caenorhabditis elegans, LON-2/glypican, a heparan sulfate proteoglycan, modulates UNC-6/netrin signaling and may do this through interactions with the UNC-40/DCC receptor. We show that developing axons misorient in the absence of LON-2/glypican when the SLT-1/slit guidance pathway is compromised and that LON-2/glypican functions in both the attractive and repulsive UNC-6/netrin pathways. We find that the core LON-2/glypican protein, lacking its heparan sulfate chains, and secreted forms of LON-2/glypican are functional in axon guidance. We also find that LON-2/glypican functions from the epidermal substrate cells to guide axons, and we provide evidence that LON-2/glypican associates with UNC-40/DCC receptor-expressing cells. We propose that LON-2/glypican acts as a modulator of UNC-40/DCC-mediated guidance to fine-tune axonal responses to UNC-6/netrin signals during migration.

No MeSH data available.


Related in: MedlinePlus