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Semaphorin6A acts as a gate keeper between the central and the peripheral nervous system.

Mauti O, Domanitskaya E, Andermatt I, Sadhu R, Stoeckli ET - Neural Dev (2007)

Bottom Line: Ablation of the boundary cap resulted in emigration of motoneurons from the ventral spinal cord along the ventral roots.Loss of either PlexinA4 or Sema6D function had an effect only at the dorsal root entry site but not at the ventral motor axon exit point.At the dorsal root entry site it organizes the segregation of dorsal roots.

View Article: PubMed Central - HTML - PubMed

Affiliation: Developmental Neuroscience, Institute of Zoology, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland. Olivier.Mauti@zool.uzh.ch

ABSTRACT

Background: During spinal cord development, expression of chicken SEMAPHORIN6A (SEMA6A) is almost exclusively found in the boundary caps at the ventral motor axon exit point and at the dorsal root entry site. The boundary cap cells are derived from a population of late migrating neural crest cells. They form a transient structure at the transition zone between the peripheral nervous system (PNS) and the central nervous system (CNS). Ablation of the boundary cap resulted in emigration of motoneurons from the ventral spinal cord along the ventral roots. Based on its very restricted expression in boundary cap cells, we tested for a role of Sema6A as a gate keeper between the CNS and the PNS.

Results: Downregulation of Sema6A in boundary cap cells by in ovo RNA interference resulted in motoneurons streaming out of the spinal cord along the ventral roots, and in the failure of dorsal roots to form and segregate properly. PlexinAs interact with class 6 semaphorins and are expressed by both motoneurons and sensory neurons. Knockdown of PlexinA1 reproduced the phenotype seen after loss of Sema6A function both at the ventral motor exit point and at the dorsal root entry site of the lumbosacral spinal cord. Loss of either PlexinA4 or Sema6D function had an effect only at the dorsal root entry site but not at the ventral motor axon exit point.

Conclusion: Sema6A acts as a gate keeper between the PNS and the CNS both ventrally and dorsally. It is required for the clustering of boundary cap cells at the PNS/CNS interface and, thus, prevents motoneurons from streaming out of the ventral spinal cord. At the dorsal root entry site it organizes the segregation of dorsal roots.

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Downregulation of PlexinA1 results in the same phenotype as seen in the absence of Sema6A. (a, b) Motoneurons streaming out of the ventral spinal cord identified by Isl-1 staining were only found after downregulation of PlexinA1 (arrows). The open arrow points to a motoneuron that is located in the ventral funiculus. Note that sensory neurons in the DRG (asterisk) are also stained by Isl-1. (e) Lack of none of the other PlexinAs enhanced the number of motoneurons found along the ventral roots compared to control-treated embryos (p = 0.0001 for the comparison between dsPA1 and all other treatment groups (indicated by three asterisks); values are given as mean ± standard error of the mean; see Figure 2b). (c, d) The phenotype seen after downregulation of Sema6A in dorsal BCCs was mimicked by both lack of PlexinA1 (c) and PlexinA4 (d). The effects of PlexinA downregulation were qualitatively different, however. In the absence of PlexinAs, the arrangement of DRGs, and not only the arrangement of their roots, was disorganized. (f) A phenotype was seen in 83% of embryos lacking PlexinA1 and in 67% of the embryos lacking PlexinA4. Bar 50 μm in (a, b); 200 μm in (c, d).
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Figure 5: Downregulation of PlexinA1 results in the same phenotype as seen in the absence of Sema6A. (a, b) Motoneurons streaming out of the ventral spinal cord identified by Isl-1 staining were only found after downregulation of PlexinA1 (arrows). The open arrow points to a motoneuron that is located in the ventral funiculus. Note that sensory neurons in the DRG (asterisk) are also stained by Isl-1. (e) Lack of none of the other PlexinAs enhanced the number of motoneurons found along the ventral roots compared to control-treated embryos (p = 0.0001 for the comparison between dsPA1 and all other treatment groups (indicated by three asterisks); values are given as mean ± standard error of the mean; see Figure 2b). (c, d) The phenotype seen after downregulation of Sema6A in dorsal BCCs was mimicked by both lack of PlexinA1 (c) and PlexinA4 (d). The effects of PlexinA downregulation were qualitatively different, however. In the absence of PlexinAs, the arrangement of DRGs, and not only the arrangement of their roots, was disorganized. (f) A phenotype was seen in 83% of embryos lacking PlexinA1 and in 67% of the embryos lacking PlexinA4. Bar 50 μm in (a, b); 200 μm in (c, d).

Mentions: We first knocked down PlexinAs in motoneurons using in ovo RNAi. For each PlexinA we used two independent long dsRNAs (see Materials and methods). Downregulation was specific for the targeted gene (Additional file 1). Consistent with its strong expression in motoneurons at the time when they extend their axons out of the VMEP, we found pronounced effects after downregulation of PlexinA1. In 34% of the sections from the lumbosacral region that we analyzed we found groups of motoneurons along the ventral root (Figure 5). All embryos lacking PlexinA1 were affected and had motoneurons outside the spinal cord in 13–52% of the sections taken from the lumbosacral spinal cord. Thus, the phenotype observed after RNAi for PLEXINA1 was qualitatively and quantitatively comparable to the phenotype observed after RNAi for SEMA6A (compare Figures 5a,b,e and 2a,c). Downregulation of PlexinA2 and A4 had no effect on the migratory behavior of motoneurons. The number of motoneurons outside the spinal cord was not different from control (Figure 5e). We counted ectopic motoneurons in 9% of the lumbosacral sections from embryos lacking PlexinA2 or PlexinA4 compared to 8% for control-treated embryos.


Semaphorin6A acts as a gate keeper between the central and the peripheral nervous system.

Mauti O, Domanitskaya E, Andermatt I, Sadhu R, Stoeckli ET - Neural Dev (2007)

Downregulation of PlexinA1 results in the same phenotype as seen in the absence of Sema6A. (a, b) Motoneurons streaming out of the ventral spinal cord identified by Isl-1 staining were only found after downregulation of PlexinA1 (arrows). The open arrow points to a motoneuron that is located in the ventral funiculus. Note that sensory neurons in the DRG (asterisk) are also stained by Isl-1. (e) Lack of none of the other PlexinAs enhanced the number of motoneurons found along the ventral roots compared to control-treated embryos (p = 0.0001 for the comparison between dsPA1 and all other treatment groups (indicated by three asterisks); values are given as mean ± standard error of the mean; see Figure 2b). (c, d) The phenotype seen after downregulation of Sema6A in dorsal BCCs was mimicked by both lack of PlexinA1 (c) and PlexinA4 (d). The effects of PlexinA downregulation were qualitatively different, however. In the absence of PlexinAs, the arrangement of DRGs, and not only the arrangement of their roots, was disorganized. (f) A phenotype was seen in 83% of embryos lacking PlexinA1 and in 67% of the embryos lacking PlexinA4. Bar 50 μm in (a, b); 200 μm in (c, d).
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Figure 5: Downregulation of PlexinA1 results in the same phenotype as seen in the absence of Sema6A. (a, b) Motoneurons streaming out of the ventral spinal cord identified by Isl-1 staining were only found after downregulation of PlexinA1 (arrows). The open arrow points to a motoneuron that is located in the ventral funiculus. Note that sensory neurons in the DRG (asterisk) are also stained by Isl-1. (e) Lack of none of the other PlexinAs enhanced the number of motoneurons found along the ventral roots compared to control-treated embryos (p = 0.0001 for the comparison between dsPA1 and all other treatment groups (indicated by three asterisks); values are given as mean ± standard error of the mean; see Figure 2b). (c, d) The phenotype seen after downregulation of Sema6A in dorsal BCCs was mimicked by both lack of PlexinA1 (c) and PlexinA4 (d). The effects of PlexinA downregulation were qualitatively different, however. In the absence of PlexinAs, the arrangement of DRGs, and not only the arrangement of their roots, was disorganized. (f) A phenotype was seen in 83% of embryos lacking PlexinA1 and in 67% of the embryos lacking PlexinA4. Bar 50 μm in (a, b); 200 μm in (c, d).
Mentions: We first knocked down PlexinAs in motoneurons using in ovo RNAi. For each PlexinA we used two independent long dsRNAs (see Materials and methods). Downregulation was specific for the targeted gene (Additional file 1). Consistent with its strong expression in motoneurons at the time when they extend their axons out of the VMEP, we found pronounced effects after downregulation of PlexinA1. In 34% of the sections from the lumbosacral region that we analyzed we found groups of motoneurons along the ventral root (Figure 5). All embryos lacking PlexinA1 were affected and had motoneurons outside the spinal cord in 13–52% of the sections taken from the lumbosacral spinal cord. Thus, the phenotype observed after RNAi for PLEXINA1 was qualitatively and quantitatively comparable to the phenotype observed after RNAi for SEMA6A (compare Figures 5a,b,e and 2a,c). Downregulation of PlexinA2 and A4 had no effect on the migratory behavior of motoneurons. The number of motoneurons outside the spinal cord was not different from control (Figure 5e). We counted ectopic motoneurons in 9% of the lumbosacral sections from embryos lacking PlexinA2 or PlexinA4 compared to 8% for control-treated embryos.

Bottom Line: Ablation of the boundary cap resulted in emigration of motoneurons from the ventral spinal cord along the ventral roots.Loss of either PlexinA4 or Sema6D function had an effect only at the dorsal root entry site but not at the ventral motor axon exit point.At the dorsal root entry site it organizes the segregation of dorsal roots.

View Article: PubMed Central - HTML - PubMed

Affiliation: Developmental Neuroscience, Institute of Zoology, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland. Olivier.Mauti@zool.uzh.ch

ABSTRACT

Background: During spinal cord development, expression of chicken SEMAPHORIN6A (SEMA6A) is almost exclusively found in the boundary caps at the ventral motor axon exit point and at the dorsal root entry site. The boundary cap cells are derived from a population of late migrating neural crest cells. They form a transient structure at the transition zone between the peripheral nervous system (PNS) and the central nervous system (CNS). Ablation of the boundary cap resulted in emigration of motoneurons from the ventral spinal cord along the ventral roots. Based on its very restricted expression in boundary cap cells, we tested for a role of Sema6A as a gate keeper between the CNS and the PNS.

Results: Downregulation of Sema6A in boundary cap cells by in ovo RNA interference resulted in motoneurons streaming out of the spinal cord along the ventral roots, and in the failure of dorsal roots to form and segregate properly. PlexinAs interact with class 6 semaphorins and are expressed by both motoneurons and sensory neurons. Knockdown of PlexinA1 reproduced the phenotype seen after loss of Sema6A function both at the ventral motor exit point and at the dorsal root entry site of the lumbosacral spinal cord. Loss of either PlexinA4 or Sema6D function had an effect only at the dorsal root entry site but not at the ventral motor axon exit point.

Conclusion: Sema6A acts as a gate keeper between the PNS and the CNS both ventrally and dorsally. It is required for the clustering of boundary cap cells at the PNS/CNS interface and, thus, prevents motoneurons from streaming out of the ventral spinal cord. At the dorsal root entry site it organizes the segregation of dorsal roots.

Show MeSH
Related in: MedlinePlus