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Ccdc80-l1 Is involved in axon pathfinding of zebrafish motoneurons.

Brusegan C, Pistocchi A, Frassine A, Della Noce I, Schepis F, Cotelli F - PLoS ONE (2012)

Bottom Line: Our results strongly suggest that ccdc80-l1 is involved in axon guidance of primary and secondary motoneurons populations, but not in their proper formation. ccdc80-l1 has a differential role as regards the development of ventral and dorsal motoneurons, and this is consistent with the asymmetric distribution of the transcript.Indeed, we reported that ccdc80-l1 expression is positively regulated by the Hedgehog pathway in adaxial cells and muscle pioneers.These findings strongly indicate ccdc80-l1 as a down-stream effector of the Hedgehog pathway.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Biologia, Università degli Studi di Milano, Milan, Italy.

ABSTRACT
Axon pathfinding is a subfield of neural development by which neurons send out axons to reach the correct targets. In particular, motoneurons extend their axons toward skeletal muscles, leading to spontaneous motor activity. In this study, we identified the zebrafish Ccdc80 and Ccdc80-like1 (Ccdc80-l1) proteins in silico on the basis of their high aminoacidic sequence identity with the human CCDC80 (Coiled-Coil Domain Containing 80). We focused on ccdc80-l1 gene that is expressed in nervous and non-nervous tissues, in particular in territories correlated with axonal migration, such as adaxial cells and muscle pioneers. Loss of ccdc80-l1 in zebrafish embryos induced motility issues, although somitogenesis and myogenesis were not impaired. Our results strongly suggest that ccdc80-l1 is involved in axon guidance of primary and secondary motoneurons populations, but not in their proper formation. ccdc80-l1 has a differential role as regards the development of ventral and dorsal motoneurons, and this is consistent with the asymmetric distribution of the transcript. The axonal migration defects observed in ccdc80-l1 loss-of-function embryos are similar to the phenotype of several mutants with altered Hedgehog activity. Indeed, we reported that ccdc80-l1 expression is positively regulated by the Hedgehog pathway in adaxial cells and muscle pioneers. These findings strongly indicate ccdc80-l1 as a down-stream effector of the Hedgehog pathway.

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Related in: MedlinePlus

Expression of ccdc80-l1 analyzed by RT-PCR and WISH.(A) RT-PCR performed on different embryonic stages and adult tissues; the expression of ccdc80-l1 and β-actin are shown. Lanes are: ladder (lane 1), ovary (lane 2), 2–4 cells stage (lane 3), 64–1000 cells stage (lane 4), 30% epiboly (lane 5), 60–70% epiboly (lane 6), somitogenesis (lane 7), 24 hpf (lane 8), 30 hpf (lane 9), 48 hpf (lane 10), 72 hpf (lane 11), adult muscle (lane 12) and negative control (lane 13) in the absence of cDNA. (B–J) WISH performed on zebrafish embryos at several stage of development. (B, C) During somitogenesis ccdc80-l1 was expressed by cranial ganglia (cg), dorsal dermis (asterisk), adaxial cells and muscle pioneers at the level of the horizontal myoseptum (arrow). (D) ccdc80-l1 expression in a transverse section of the trunk of an embryo at 12 somites stage (arrows). (E–H) At 24 hpf, the hybridization signal was detectable in cranial ganglia (cg), dermis (asterisk), adaxial cells (arrow) and ventral somites (arrowhead). (F) Higher magnification of the tail at 24 hpf. (G) Transversal section of an embryo at 24 hpf. (H) Transversal section showing that at 24 hpf ccdc80-l1 hydridization signal co-localized with the nuclear labeling of 4D9 antibody, corresponding to the engrailed-positive muscle pioneers population (open arrowhead). (I, J) At 36 hpf, the signal of ccdc80-l1 probe was detected in cranial ganglia (cg), migrated adaxial cells (arrow), dorsal dermis (asterisk) and caudal vein plexus region (cvp). (K, L) At 48 hpf, ccdc80-l1 was detected in dorsal dermis (asterisk), external adaxial cells (arrows in K) and caudal vein plexus region (cvp in L). (B, E, F, I) Lateral views; dorsal is up, anterior is left; (C) dorsal view, anterior is left; (D,G, H, J–L) transversal sections, dorsal is up.
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pone-0031851-g002: Expression of ccdc80-l1 analyzed by RT-PCR and WISH.(A) RT-PCR performed on different embryonic stages and adult tissues; the expression of ccdc80-l1 and β-actin are shown. Lanes are: ladder (lane 1), ovary (lane 2), 2–4 cells stage (lane 3), 64–1000 cells stage (lane 4), 30% epiboly (lane 5), 60–70% epiboly (lane 6), somitogenesis (lane 7), 24 hpf (lane 8), 30 hpf (lane 9), 48 hpf (lane 10), 72 hpf (lane 11), adult muscle (lane 12) and negative control (lane 13) in the absence of cDNA. (B–J) WISH performed on zebrafish embryos at several stage of development. (B, C) During somitogenesis ccdc80-l1 was expressed by cranial ganglia (cg), dorsal dermis (asterisk), adaxial cells and muscle pioneers at the level of the horizontal myoseptum (arrow). (D) ccdc80-l1 expression in a transverse section of the trunk of an embryo at 12 somites stage (arrows). (E–H) At 24 hpf, the hybridization signal was detectable in cranial ganglia (cg), dermis (asterisk), adaxial cells (arrow) and ventral somites (arrowhead). (F) Higher magnification of the tail at 24 hpf. (G) Transversal section of an embryo at 24 hpf. (H) Transversal section showing that at 24 hpf ccdc80-l1 hydridization signal co-localized with the nuclear labeling of 4D9 antibody, corresponding to the engrailed-positive muscle pioneers population (open arrowhead). (I, J) At 36 hpf, the signal of ccdc80-l1 probe was detected in cranial ganglia (cg), migrated adaxial cells (arrow), dorsal dermis (asterisk) and caudal vein plexus region (cvp). (K, L) At 48 hpf, ccdc80-l1 was detected in dorsal dermis (asterisk), external adaxial cells (arrows in K) and caudal vein plexus region (cvp in L). (B, E, F, I) Lateral views; dorsal is up, anterior is left; (C) dorsal view, anterior is left; (D,G, H, J–L) transversal sections, dorsal is up.

Mentions: Characterization of ccdc80-l1 expression, using RT-PCR, revealed that ccdc80-l1 transcript is present from the first stages of development up to 72 hpf, thus including maternal and zygotic transcription (Fig. 2A). ccdc80-l1 is also expressed in the ovary and muscle of the adult zebrafish (Fig. 2A). During somitogenesis, the hybridization signal is restricted to the horizontal myoseptum (Fig. 2B–D). From this stage, ccdc80-l1 expression is observed also in the cranial ganglia and dorsal dermis (Fig. 2B, 2C, 2E, 2I, 2K–L and data not shown). At 24 hpf, ccdc80-l1 is detectable in a specific sub-population of migrating adaxial cells, that moves along the lateral axis towards the external somite [35] (Fig. 2E–G). Moreover, ccdc80-l1 is expressed in muscle pioneers, as shown by the co-localization between ccdc80-l1 and engrailed [36], [37] (Fig. 2H). ccdc80-l1 expression in adaxial cells persisted at 36 hpf and 48 hpf (Fig. 2I, 2K). At the same stages ccdc80-l1 is also expressed in the caudal vein plexus region (Fig. 2I, 2J, 2L).


Ccdc80-l1 Is involved in axon pathfinding of zebrafish motoneurons.

Brusegan C, Pistocchi A, Frassine A, Della Noce I, Schepis F, Cotelli F - PLoS ONE (2012)

Expression of ccdc80-l1 analyzed by RT-PCR and WISH.(A) RT-PCR performed on different embryonic stages and adult tissues; the expression of ccdc80-l1 and β-actin are shown. Lanes are: ladder (lane 1), ovary (lane 2), 2–4 cells stage (lane 3), 64–1000 cells stage (lane 4), 30% epiboly (lane 5), 60–70% epiboly (lane 6), somitogenesis (lane 7), 24 hpf (lane 8), 30 hpf (lane 9), 48 hpf (lane 10), 72 hpf (lane 11), adult muscle (lane 12) and negative control (lane 13) in the absence of cDNA. (B–J) WISH performed on zebrafish embryos at several stage of development. (B, C) During somitogenesis ccdc80-l1 was expressed by cranial ganglia (cg), dorsal dermis (asterisk), adaxial cells and muscle pioneers at the level of the horizontal myoseptum (arrow). (D) ccdc80-l1 expression in a transverse section of the trunk of an embryo at 12 somites stage (arrows). (E–H) At 24 hpf, the hybridization signal was detectable in cranial ganglia (cg), dermis (asterisk), adaxial cells (arrow) and ventral somites (arrowhead). (F) Higher magnification of the tail at 24 hpf. (G) Transversal section of an embryo at 24 hpf. (H) Transversal section showing that at 24 hpf ccdc80-l1 hydridization signal co-localized with the nuclear labeling of 4D9 antibody, corresponding to the engrailed-positive muscle pioneers population (open arrowhead). (I, J) At 36 hpf, the signal of ccdc80-l1 probe was detected in cranial ganglia (cg), migrated adaxial cells (arrow), dorsal dermis (asterisk) and caudal vein plexus region (cvp). (K, L) At 48 hpf, ccdc80-l1 was detected in dorsal dermis (asterisk), external adaxial cells (arrows in K) and caudal vein plexus region (cvp in L). (B, E, F, I) Lateral views; dorsal is up, anterior is left; (C) dorsal view, anterior is left; (D,G, H, J–L) transversal sections, dorsal is up.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3285184&req=5

pone-0031851-g002: Expression of ccdc80-l1 analyzed by RT-PCR and WISH.(A) RT-PCR performed on different embryonic stages and adult tissues; the expression of ccdc80-l1 and β-actin are shown. Lanes are: ladder (lane 1), ovary (lane 2), 2–4 cells stage (lane 3), 64–1000 cells stage (lane 4), 30% epiboly (lane 5), 60–70% epiboly (lane 6), somitogenesis (lane 7), 24 hpf (lane 8), 30 hpf (lane 9), 48 hpf (lane 10), 72 hpf (lane 11), adult muscle (lane 12) and negative control (lane 13) in the absence of cDNA. (B–J) WISH performed on zebrafish embryos at several stage of development. (B, C) During somitogenesis ccdc80-l1 was expressed by cranial ganglia (cg), dorsal dermis (asterisk), adaxial cells and muscle pioneers at the level of the horizontal myoseptum (arrow). (D) ccdc80-l1 expression in a transverse section of the trunk of an embryo at 12 somites stage (arrows). (E–H) At 24 hpf, the hybridization signal was detectable in cranial ganglia (cg), dermis (asterisk), adaxial cells (arrow) and ventral somites (arrowhead). (F) Higher magnification of the tail at 24 hpf. (G) Transversal section of an embryo at 24 hpf. (H) Transversal section showing that at 24 hpf ccdc80-l1 hydridization signal co-localized with the nuclear labeling of 4D9 antibody, corresponding to the engrailed-positive muscle pioneers population (open arrowhead). (I, J) At 36 hpf, the signal of ccdc80-l1 probe was detected in cranial ganglia (cg), migrated adaxial cells (arrow), dorsal dermis (asterisk) and caudal vein plexus region (cvp). (K, L) At 48 hpf, ccdc80-l1 was detected in dorsal dermis (asterisk), external adaxial cells (arrows in K) and caudal vein plexus region (cvp in L). (B, E, F, I) Lateral views; dorsal is up, anterior is left; (C) dorsal view, anterior is left; (D,G, H, J–L) transversal sections, dorsal is up.
Mentions: Characterization of ccdc80-l1 expression, using RT-PCR, revealed that ccdc80-l1 transcript is present from the first stages of development up to 72 hpf, thus including maternal and zygotic transcription (Fig. 2A). ccdc80-l1 is also expressed in the ovary and muscle of the adult zebrafish (Fig. 2A). During somitogenesis, the hybridization signal is restricted to the horizontal myoseptum (Fig. 2B–D). From this stage, ccdc80-l1 expression is observed also in the cranial ganglia and dorsal dermis (Fig. 2B, 2C, 2E, 2I, 2K–L and data not shown). At 24 hpf, ccdc80-l1 is detectable in a specific sub-population of migrating adaxial cells, that moves along the lateral axis towards the external somite [35] (Fig. 2E–G). Moreover, ccdc80-l1 is expressed in muscle pioneers, as shown by the co-localization between ccdc80-l1 and engrailed [36], [37] (Fig. 2H). ccdc80-l1 expression in adaxial cells persisted at 36 hpf and 48 hpf (Fig. 2I, 2K). At the same stages ccdc80-l1 is also expressed in the caudal vein plexus region (Fig. 2I, 2J, 2L).

Bottom Line: Our results strongly suggest that ccdc80-l1 is involved in axon guidance of primary and secondary motoneurons populations, but not in their proper formation. ccdc80-l1 has a differential role as regards the development of ventral and dorsal motoneurons, and this is consistent with the asymmetric distribution of the transcript.Indeed, we reported that ccdc80-l1 expression is positively regulated by the Hedgehog pathway in adaxial cells and muscle pioneers.These findings strongly indicate ccdc80-l1 as a down-stream effector of the Hedgehog pathway.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Biologia, Università degli Studi di Milano, Milan, Italy.

ABSTRACT
Axon pathfinding is a subfield of neural development by which neurons send out axons to reach the correct targets. In particular, motoneurons extend their axons toward skeletal muscles, leading to spontaneous motor activity. In this study, we identified the zebrafish Ccdc80 and Ccdc80-like1 (Ccdc80-l1) proteins in silico on the basis of their high aminoacidic sequence identity with the human CCDC80 (Coiled-Coil Domain Containing 80). We focused on ccdc80-l1 gene that is expressed in nervous and non-nervous tissues, in particular in territories correlated with axonal migration, such as adaxial cells and muscle pioneers. Loss of ccdc80-l1 in zebrafish embryos induced motility issues, although somitogenesis and myogenesis were not impaired. Our results strongly suggest that ccdc80-l1 is involved in axon guidance of primary and secondary motoneurons populations, but not in their proper formation. ccdc80-l1 has a differential role as regards the development of ventral and dorsal motoneurons, and this is consistent with the asymmetric distribution of the transcript. The axonal migration defects observed in ccdc80-l1 loss-of-function embryos are similar to the phenotype of several mutants with altered Hedgehog activity. Indeed, we reported that ccdc80-l1 expression is positively regulated by the Hedgehog pathway in adaxial cells and muscle pioneers. These findings strongly indicate ccdc80-l1 as a down-stream effector of the Hedgehog pathway.

Show MeSH
Related in: MedlinePlus