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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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Axons of DRG and sympathetic neurons but not motor axons are repelled by Sema6A. (a) Upon encountering a COS cell expressing Sema6A, 53% of all DRG axons were found to react with avoidance, that is, they turned away from the cell or stopped rather than growing onto the Sema6A-positive COS cell (p = 0.0002 for the comparison between S6A/Ax-1 and S6A/EGFP (indicated by three asterisks)). COS cells expressing Axonin-1 (Ax-1) were perceived as slightly more attractive than control COS cells expressing only EGFP (p = 0.006 for S6A/Ax-1 and 0.02 for S6A/EGFP (indicated by asterisk)). (b) Motor axons were found to be indifferent to all types of COS cells. The majority showed neither attraction nor repulsion when encountering Sema6A or Axonin-1 compared to EGFP-expressing COS cells. (c) The majority of sympathetic axons (68%) were avoiding Sema6A-expressing cells (p < 0.0001 (three asterisks)). Compared to DRG axons and motor axons, sympathetic axons were more strongly attracted by Axonin-1-expressing COS cells (p < 0.0001 for Ax-1/S6A and Ax-1/EGFP). This is reflected by the fact that avoidance of Axonin-1-expressing cells was significantly lower compared to EGFP-expressing cells (p = 0.003 (two asterisks)). Similarly, axons of sympathetic neurons (SG) were significantly less attracted to Sema6A-expressing compared to EGFP-expressing cells (p < 0.0001 (three asterisks)). Values are given as mean ± standard error of the mean.
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Figure 6: Axons of DRG and sympathetic neurons but not motor axons are repelled by Sema6A. (a) Upon encountering a COS cell expressing Sema6A, 53% of all DRG axons were found to react with avoidance, that is, they turned away from the cell or stopped rather than growing onto the Sema6A-positive COS cell (p = 0.0002 for the comparison between S6A/Ax-1 and S6A/EGFP (indicated by three asterisks)). COS cells expressing Axonin-1 (Ax-1) were perceived as slightly more attractive than control COS cells expressing only EGFP (p = 0.006 for S6A/Ax-1 and 0.02 for S6A/EGFP (indicated by asterisk)). (b) Motor axons were found to be indifferent to all types of COS cells. The majority showed neither attraction nor repulsion when encountering Sema6A or Axonin-1 compared to EGFP-expressing COS cells. (c) The majority of sympathetic axons (68%) were avoiding Sema6A-expressing cells (p < 0.0001 (three asterisks)). Compared to DRG axons and motor axons, sympathetic axons were more strongly attracted by Axonin-1-expressing COS cells (p < 0.0001 for Ax-1/S6A and Ax-1/EGFP). This is reflected by the fact that avoidance of Axonin-1-expressing cells was significantly lower compared to EGFP-expressing cells (p = 0.003 (two asterisks)). Similarly, axons of sympathetic neurons (SG) were significantly less attracted to Sema6A-expressing compared to EGFP-expressing cells (p < 0.0001 (three asterisks)). Values are given as mean ± standard error of the mean.

Mentions: To get a lead on the mechanism of Sema6A function in boundary control, we turned to an in vitro assay (Figure 6). We wanted to assess whether Sema6A in BCCs had an attractive or a repulsive effect on sensory and motor axons, respectively. For this purpose, we transfected COS cells with SEMA6A and used them as a substrate for DRG neurons and motoneurons. We also used sympathetic neurons as they were shown to react to Sema6A contact with growth cone collapse [34]. Axonin-1 was used as a control protein. We scored the behavior of axons encountering transfected COS cells as 'repulsion' when axon failed to grow onto a transfected cell by either stopping or turning away. The score was 'attraction' when axons readily crossed from a non-transfected to a transfected COS cell but did not cross back from the transfected to a non-transfected cell. Axons that readily crossed from a non-transfected to a transfected cell and back to a non-transfected cell were scored as 'crossing', or, in other words, were considered not to be affected by the protein expressed on COS cells. COS cells expressing EGFP were used as an additional control to measure the 'baseline behavior' of axons growing on COS cells. As expected, few cells reacted with repulsion or attraction to COS cells expressing EGFP. For all types of neurons, we found that more than 70% of the axons crossed EGFP-transfected COS cells readily (Table 1). The behavior was different in response to COS cells expressing Sema6A. Both DRG (Figure 6a) and sympathetic axons (Figure 6c) avoided Sema6A-positive cells. The effect was stronger for sympathetic neurons, where avoidance was found for 68% of the axons compared to 53% of the DRG axons (Table 1). Axons of motoneurons did not show a reaction to either Sema6A or Axonin-1 that differed from the behavior on EGFP-expressing cells (Figure 6b). Interestingly, we found that significantly more sympathetic axons reacted with attraction to Axonin-1 than to control COS cells expressing EGFP (Figure 6c). In conclusion, axons of DRG neurons were repelled by Sema6A, whereas motor axons did not react at all to Sema6A.


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)

Axons of DRG and sympathetic neurons but not motor axons are repelled by Sema6A. (a) Upon encountering a COS cell expressing Sema6A, 53% of all DRG axons were found to react with avoidance, that is, they turned away from the cell or stopped rather than growing onto the Sema6A-positive COS cell (p = 0.0002 for the comparison between S6A/Ax-1 and S6A/EGFP (indicated by three asterisks)). COS cells expressing Axonin-1 (Ax-1) were perceived as slightly more attractive than control COS cells expressing only EGFP (p = 0.006 for S6A/Ax-1 and 0.02 for S6A/EGFP (indicated by asterisk)). (b) Motor axons were found to be indifferent to all types of COS cells. The majority showed neither attraction nor repulsion when encountering Sema6A or Axonin-1 compared to EGFP-expressing COS cells. (c) The majority of sympathetic axons (68%) were avoiding Sema6A-expressing cells (p < 0.0001 (three asterisks)). Compared to DRG axons and motor axons, sympathetic axons were more strongly attracted by Axonin-1-expressing COS cells (p < 0.0001 for Ax-1/S6A and Ax-1/EGFP). This is reflected by the fact that avoidance of Axonin-1-expressing cells was significantly lower compared to EGFP-expressing cells (p = 0.003 (two asterisks)). Similarly, axons of sympathetic neurons (SG) were significantly less attracted to Sema6A-expressing compared to EGFP-expressing cells (p < 0.0001 (three asterisks)). Values are given as mean ± standard error of the mean.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 6: Axons of DRG and sympathetic neurons but not motor axons are repelled by Sema6A. (a) Upon encountering a COS cell expressing Sema6A, 53% of all DRG axons were found to react with avoidance, that is, they turned away from the cell or stopped rather than growing onto the Sema6A-positive COS cell (p = 0.0002 for the comparison between S6A/Ax-1 and S6A/EGFP (indicated by three asterisks)). COS cells expressing Axonin-1 (Ax-1) were perceived as slightly more attractive than control COS cells expressing only EGFP (p = 0.006 for S6A/Ax-1 and 0.02 for S6A/EGFP (indicated by asterisk)). (b) Motor axons were found to be indifferent to all types of COS cells. The majority showed neither attraction nor repulsion when encountering Sema6A or Axonin-1 compared to EGFP-expressing COS cells. (c) The majority of sympathetic axons (68%) were avoiding Sema6A-expressing cells (p < 0.0001 (three asterisks)). Compared to DRG axons and motor axons, sympathetic axons were more strongly attracted by Axonin-1-expressing COS cells (p < 0.0001 for Ax-1/S6A and Ax-1/EGFP). This is reflected by the fact that avoidance of Axonin-1-expressing cells was significantly lower compared to EGFP-expressing cells (p = 0.003 (two asterisks)). Similarly, axons of sympathetic neurons (SG) were significantly less attracted to Sema6A-expressing compared to EGFP-expressing cells (p < 0.0001 (three asterisks)). Values are given as mean ± standard error of the mean.
Mentions: To get a lead on the mechanism of Sema6A function in boundary control, we turned to an in vitro assay (Figure 6). We wanted to assess whether Sema6A in BCCs had an attractive or a repulsive effect on sensory and motor axons, respectively. For this purpose, we transfected COS cells with SEMA6A and used them as a substrate for DRG neurons and motoneurons. We also used sympathetic neurons as they were shown to react to Sema6A contact with growth cone collapse [34]. Axonin-1 was used as a control protein. We scored the behavior of axons encountering transfected COS cells as 'repulsion' when axon failed to grow onto a transfected cell by either stopping or turning away. The score was 'attraction' when axons readily crossed from a non-transfected to a transfected COS cell but did not cross back from the transfected to a non-transfected cell. Axons that readily crossed from a non-transfected to a transfected cell and back to a non-transfected cell were scored as 'crossing', or, in other words, were considered not to be affected by the protein expressed on COS cells. COS cells expressing EGFP were used as an additional control to measure the 'baseline behavior' of axons growing on COS cells. As expected, few cells reacted with repulsion or attraction to COS cells expressing EGFP. For all types of neurons, we found that more than 70% of the axons crossed EGFP-transfected COS cells readily (Table 1). The behavior was different in response to COS cells expressing Sema6A. Both DRG (Figure 6a) and sympathetic axons (Figure 6c) avoided Sema6A-positive cells. The effect was stronger for sympathetic neurons, where avoidance was found for 68% of the axons compared to 53% of the DRG axons (Table 1). Axons of motoneurons did not show a reaction to either Sema6A or Axonin-1 that differed from the behavior on EGFP-expressing cells (Figure 6b). Interestingly, we found that significantly more sympathetic axons reacted with attraction to Axonin-1 than to control COS cells expressing EGFP (Figure 6c). In conclusion, axons of DRG neurons were repelled by Sema6A, whereas motor axons did not react at all to Sema6A.

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