Limits...
NOVA regulates Dcc alternative splicing during neuronal migration and axon guidance in the spinal cord.

Leggere JC, Saito Y, Darnell RB, Tessier-Lavigne M, Junge HJ, Chen Z - Elife (2016)

Bottom Line: RNA-binding proteins (RBPs) control multiple aspects of post-transcriptional gene regulation and function during various biological processes in the nervous system.We found that the NOVA family of RBPs play a key role in neuronal migration, axon outgrowth, and axon guidance.Together, our results demonstrate that the production of DCC splice variants controlled by NOVA has a crucial function during many stages of commissural neuron development.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, United States.

ABSTRACT
RNA-binding proteins (RBPs) control multiple aspects of post-transcriptional gene regulation and function during various biological processes in the nervous system. To further reveal the functional significance of RBPs during neural development, we carried out an in vivo RNAi screen in the dorsal spinal cord interneurons, including the commissural neurons. We found that the NOVA family of RBPs play a key role in neuronal migration, axon outgrowth, and axon guidance. Interestingly, Nova mutants display similar defects as the knockout of the Dcc transmembrane receptor. We show here that Nova deficiency disrupts the alternative splicing of Dcc, and that restoring Dcc splicing in Nova knockouts is able to rescue the defects. Together, our results demonstrate that the production of DCC splice variants controlled by NOVA has a crucial function during many stages of commissural neuron development.

No MeSH data available.


Related in: MedlinePlus

Neural stem cells and progenitors appear normal in Nova and Dcc KOs.(A) Immunohistochemistry of phospho-histone H3, a mitotic marker, Ki-67, a cell proliferation marker, SOX2, a neural progenitor marker, and PAX3/7, a dorsal interneuron progenitor marker, in E10.5 spinal cords. (B) Quantification of phenotypes in A. pH3+ cells were counted and normalized to WT. The ratio between the SOX2+ VZ area and the total spinal cord area was quantified and normalized to WT. Data are represented as the mean ± SEM (Student’s t-test, ns, not significant). Scale bar, 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14264.011
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4930329&req=5

fig3s1: Neural stem cells and progenitors appear normal in Nova and Dcc KOs.(A) Immunohistochemistry of phospho-histone H3, a mitotic marker, Ki-67, a cell proliferation marker, SOX2, a neural progenitor marker, and PAX3/7, a dorsal interneuron progenitor marker, in E10.5 spinal cords. (B) Quantification of phenotypes in A. pH3+ cells were counted and normalized to WT. The ratio between the SOX2+ VZ area and the total spinal cord area was quantified and normalized to WT. Data are represented as the mean ± SEM (Student’s t-test, ns, not significant). Scale bar, 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14264.011

Mentions: The increase in GFP+ neuroprogenitors in the VZ could result from slowed migration, increased proliferation, or reduced neuronal differentiation. To follow cell migration, we electroporated the progenitors with Actb-gfp and cultured the embryos for 20 hrs. In WT embryos, we observed that many progenitors have reached the lateral spinal cord after 20 hrs. However, in both Nova and Dcc KOs, almost all GFP+ neurons are still positioned within the VZ (Figure 3). This supports the idea that neuronal migration is reduced. We examined cell proliferation in Nova and Dcc KOs using cell cycle markers, including phospho-histone H3 (pH3), a mitotic marker, and Ki-67, a cell proliferation marker. We found that at E10.5, the number of neural stem cells and progenitors is normal in both mutants (Figure 3—figure supplement 1). In addition, we labeled the neural progenitors in the whole spinal cord with SOX2 and the dorsal progenitors with PAX3/7, and found no change in the localization or organization of these progenitor populations (Figure 3—figure supplement 1). We also examined interneuron differentiation using the BARHL2, ISL1/2, and LHX5 markers, and found that a normal number of interneurons are born in Nova and Dcc KOs at E10.5 (Figure 3—figure supplement 2; Figure 4). Therefore, both Nova and Dcc deficiency reduces neuroprogenitor migration, but does not affect cell proliferation or interneuron differentiation.10.7554/eLife.14264.010Figure 3.Nova and Dcc knockouts delay the lateral migration of dorsal interneuron progenitors.


NOVA regulates Dcc alternative splicing during neuronal migration and axon guidance in the spinal cord.

Leggere JC, Saito Y, Darnell RB, Tessier-Lavigne M, Junge HJ, Chen Z - Elife (2016)

Neural stem cells and progenitors appear normal in Nova and Dcc KOs.(A) Immunohistochemistry of phospho-histone H3, a mitotic marker, Ki-67, a cell proliferation marker, SOX2, a neural progenitor marker, and PAX3/7, a dorsal interneuron progenitor marker, in E10.5 spinal cords. (B) Quantification of phenotypes in A. pH3+ cells were counted and normalized to WT. The ratio between the SOX2+ VZ area and the total spinal cord area was quantified and normalized to WT. Data are represented as the mean ± SEM (Student’s t-test, ns, not significant). Scale bar, 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14264.011
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4930329&req=5

fig3s1: Neural stem cells and progenitors appear normal in Nova and Dcc KOs.(A) Immunohistochemistry of phospho-histone H3, a mitotic marker, Ki-67, a cell proliferation marker, SOX2, a neural progenitor marker, and PAX3/7, a dorsal interneuron progenitor marker, in E10.5 spinal cords. (B) Quantification of phenotypes in A. pH3+ cells were counted and normalized to WT. The ratio between the SOX2+ VZ area and the total spinal cord area was quantified and normalized to WT. Data are represented as the mean ± SEM (Student’s t-test, ns, not significant). Scale bar, 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14264.011
Mentions: The increase in GFP+ neuroprogenitors in the VZ could result from slowed migration, increased proliferation, or reduced neuronal differentiation. To follow cell migration, we electroporated the progenitors with Actb-gfp and cultured the embryos for 20 hrs. In WT embryos, we observed that many progenitors have reached the lateral spinal cord after 20 hrs. However, in both Nova and Dcc KOs, almost all GFP+ neurons are still positioned within the VZ (Figure 3). This supports the idea that neuronal migration is reduced. We examined cell proliferation in Nova and Dcc KOs using cell cycle markers, including phospho-histone H3 (pH3), a mitotic marker, and Ki-67, a cell proliferation marker. We found that at E10.5, the number of neural stem cells and progenitors is normal in both mutants (Figure 3—figure supplement 1). In addition, we labeled the neural progenitors in the whole spinal cord with SOX2 and the dorsal progenitors with PAX3/7, and found no change in the localization or organization of these progenitor populations (Figure 3—figure supplement 1). We also examined interneuron differentiation using the BARHL2, ISL1/2, and LHX5 markers, and found that a normal number of interneurons are born in Nova and Dcc KOs at E10.5 (Figure 3—figure supplement 2; Figure 4). Therefore, both Nova and Dcc deficiency reduces neuroprogenitor migration, but does not affect cell proliferation or interneuron differentiation.10.7554/eLife.14264.010Figure 3.Nova and Dcc knockouts delay the lateral migration of dorsal interneuron progenitors.

Bottom Line: RNA-binding proteins (RBPs) control multiple aspects of post-transcriptional gene regulation and function during various biological processes in the nervous system.We found that the NOVA family of RBPs play a key role in neuronal migration, axon outgrowth, and axon guidance.Together, our results demonstrate that the production of DCC splice variants controlled by NOVA has a crucial function during many stages of commissural neuron development.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, United States.

ABSTRACT
RNA-binding proteins (RBPs) control multiple aspects of post-transcriptional gene regulation and function during various biological processes in the nervous system. To further reveal the functional significance of RBPs during neural development, we carried out an in vivo RNAi screen in the dorsal spinal cord interneurons, including the commissural neurons. We found that the NOVA family of RBPs play a key role in neuronal migration, axon outgrowth, and axon guidance. Interestingly, Nova mutants display similar defects as the knockout of the Dcc transmembrane receptor. We show here that Nova deficiency disrupts the alternative splicing of Dcc, and that restoring Dcc splicing in Nova knockouts is able to rescue the defects. Together, our results demonstrate that the production of DCC splice variants controlled by NOVA has a crucial function during many stages of commissural neuron development.

No MeSH data available.


Related in: MedlinePlus