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NOVA2-mediated RNA regulation is required for axonal pathfinding during development.

Saito Y, Miranda-Rottmann S, Ruggiu M, Park CY, Fak JJ, Zhong R, Duncan JS, Fabella BA, Junge HJ, Chen Z, Araya R, Fritzsch B, Hudspeth AJ, Darnell RB - Elife (2016)

Bottom Line: The neuron specific RNA-binding proteins NOVA1 and NOVA2 are highly homologous alternative splicing regulators.NOVA proteins regulate at least 700 alternative splicing events in vivo, yet relatively little is known about the biologic consequences of NOVA action and in particular about functional differences between NOVA1 and NOVA2.Thus we have discovered that NOVA2 uniquely regulates alternative splicing of a coordinate set of transcripts encoding key components in cortical, brainstem and spinal axon guidance/outgrowth pathways during neural differentiation, with severe functional consequences in vivo.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Neuro-Oncology, Howard Hughes Medical Institute, The Rockefeller University, New York, United States.

ABSTRACT
The neuron specific RNA-binding proteins NOVA1 and NOVA2 are highly homologous alternative splicing regulators. NOVA proteins regulate at least 700 alternative splicing events in vivo, yet relatively little is known about the biologic consequences of NOVA action and in particular about functional differences between NOVA1 and NOVA2. Transcriptome-wide searches for isoform-specific functions, using NOVA1 and NOVA2 specific HITS-CLIP and RNA-seq data from mouse cortex lacking either NOVA isoform, reveals that NOVA2 uniquely regulates alternative splicing events of a series of axon guidance related genes during cortical development. Corresponding axonal pathfinding defects were specific to NOVA2 deficiency: Nova2-/- but not Nova1-/- mice had agenesis of the corpus callosum, and axonal outgrowth defects specific to ventral motoneuron axons and efferent innervation of the cochlea. Thus we have discovered that NOVA2 uniquely regulates alternative splicing of a coordinate set of transcripts encoding key components in cortical, brainstem and spinal axon guidance/outgrowth pathways during neural differentiation, with severe functional consequences in vivo.

No MeSH data available.


Related in: MedlinePlus

Abnormal thalamo-cortical path in the cortex of Nova2-/- mice.(A) Immunohistochemistry of L1 (c,d) and NOVA2 (e,f) on coronal sections in wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NOVA2 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Aa) and (Ab). Scale bars; 200 μm (a,b), 50 μm (c–h). (B) Immunohistochemistry of L1 (c,d) and NTNG1 (e,f) on coronal sections in wild-type (a, c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NTNG1 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Ba) and (Bb). Scale bars; 500 μm (a,b), 50 μm (c–h). (C) Immunohistochemistry of L1 (c,d) and NURR1 (e,f) in the cortex of wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged views of L1 (green), NURR1 (red), and DAPI (blue). Scale bar; 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14371.024
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fig7: Abnormal thalamo-cortical path in the cortex of Nova2-/- mice.(A) Immunohistochemistry of L1 (c,d) and NOVA2 (e,f) on coronal sections in wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NOVA2 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Aa) and (Ab). Scale bars; 200 μm (a,b), 50 μm (c–h). (B) Immunohistochemistry of L1 (c,d) and NTNG1 (e,f) on coronal sections in wild-type (a, c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NTNG1 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Ba) and (Bb). Scale bars; 500 μm (a,b), 50 μm (c–h). (C) Immunohistochemistry of L1 (c,d) and NURR1 (e,f) in the cortex of wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged views of L1 (green), NURR1 (red), and DAPI (blue). Scale bar; 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14371.024

Mentions: To understand the axon guidance phenotype in more detail, we compared the pathway of L1 positive axons traversing the upper cortical layer in the cortex of Nova2-/- and wild-type mice at E16.5 and E18.5 (E16.5; Figure 7 and E18.5; Figure 6A). In wild-type mice, L1 positive axons passing between the subplate and subventricular zone were detected as one wide bundle, whereas in Nova2-/- mice L1 positive axon routes were separated into two bundles (Figure 7A). The L1 positive axons detected in the deeper cortical layer in Nova2-/- appeared normal, in that they passed through similar cortical layers as did axons in wild-type mice. In contrast, separated axons passing through the upper L1 positive axonal pathway, which were Netrin-G1 (NTNG1) positive axons, were only seen in Nova2-/- mice and were passed along a subplate path enriched in NURR1 (a subplate marker), as revealed by L1/NTNG1 or L1/NURR1 double-immunostaining (Figure 7B and Figure 7C). These results indicate that only a portion of the L1 positive axonal path, which is NTNG1 positive, is specifically affected by the absence of Nova2. We conclude that NOVA2 plays a role as a modifier for a unique set of neuron-specific axons in the developing cortex.10.7554/eLife.14371.024Figure 7.Abnormal thalamo-cortical path in the cortex of Nova2-/- mice.


NOVA2-mediated RNA regulation is required for axonal pathfinding during development.

Saito Y, Miranda-Rottmann S, Ruggiu M, Park CY, Fak JJ, Zhong R, Duncan JS, Fabella BA, Junge HJ, Chen Z, Araya R, Fritzsch B, Hudspeth AJ, Darnell RB - Elife (2016)

Abnormal thalamo-cortical path in the cortex of Nova2-/- mice.(A) Immunohistochemistry of L1 (c,d) and NOVA2 (e,f) on coronal sections in wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NOVA2 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Aa) and (Ab). Scale bars; 200 μm (a,b), 50 μm (c–h). (B) Immunohistochemistry of L1 (c,d) and NTNG1 (e,f) on coronal sections in wild-type (a, c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NTNG1 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Ba) and (Bb). Scale bars; 500 μm (a,b), 50 μm (c–h). (C) Immunohistochemistry of L1 (c,d) and NURR1 (e,f) in the cortex of wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged views of L1 (green), NURR1 (red), and DAPI (blue). Scale bar; 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14371.024
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fig7: Abnormal thalamo-cortical path in the cortex of Nova2-/- mice.(A) Immunohistochemistry of L1 (c,d) and NOVA2 (e,f) on coronal sections in wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NOVA2 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Aa) and (Ab). Scale bars; 200 μm (a,b), 50 μm (c–h). (B) Immunohistochemistry of L1 (c,d) and NTNG1 (e,f) on coronal sections in wild-type (a, c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged coronal section views of L1 (green), NTNG1 (red), and DAPI (blue). (c-h) Higher magnified view of neocortex of (Ba) and (Bb). Scale bars; 500 μm (a,b), 50 μm (c–h). (C) Immunohistochemistry of L1 (c,d) and NURR1 (e,f) in the cortex of wild-type (a,c,e,g) and Nova2-/- (b,d,f,h) at E16.5. (a,b) Merged views of L1 (green), NURR1 (red), and DAPI (blue). Scale bar; 50 μm.DOI:http://dx.doi.org/10.7554/eLife.14371.024
Mentions: To understand the axon guidance phenotype in more detail, we compared the pathway of L1 positive axons traversing the upper cortical layer in the cortex of Nova2-/- and wild-type mice at E16.5 and E18.5 (E16.5; Figure 7 and E18.5; Figure 6A). In wild-type mice, L1 positive axons passing between the subplate and subventricular zone were detected as one wide bundle, whereas in Nova2-/- mice L1 positive axon routes were separated into two bundles (Figure 7A). The L1 positive axons detected in the deeper cortical layer in Nova2-/- appeared normal, in that they passed through similar cortical layers as did axons in wild-type mice. In contrast, separated axons passing through the upper L1 positive axonal pathway, which were Netrin-G1 (NTNG1) positive axons, were only seen in Nova2-/- mice and were passed along a subplate path enriched in NURR1 (a subplate marker), as revealed by L1/NTNG1 or L1/NURR1 double-immunostaining (Figure 7B and Figure 7C). These results indicate that only a portion of the L1 positive axonal path, which is NTNG1 positive, is specifically affected by the absence of Nova2. We conclude that NOVA2 plays a role as a modifier for a unique set of neuron-specific axons in the developing cortex.10.7554/eLife.14371.024Figure 7.Abnormal thalamo-cortical path in the cortex of Nova2-/- mice.

Bottom Line: The neuron specific RNA-binding proteins NOVA1 and NOVA2 are highly homologous alternative splicing regulators.NOVA proteins regulate at least 700 alternative splicing events in vivo, yet relatively little is known about the biologic consequences of NOVA action and in particular about functional differences between NOVA1 and NOVA2.Thus we have discovered that NOVA2 uniquely regulates alternative splicing of a coordinate set of transcripts encoding key components in cortical, brainstem and spinal axon guidance/outgrowth pathways during neural differentiation, with severe functional consequences in vivo.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Neuro-Oncology, Howard Hughes Medical Institute, The Rockefeller University, New York, United States.

ABSTRACT
The neuron specific RNA-binding proteins NOVA1 and NOVA2 are highly homologous alternative splicing regulators. NOVA proteins regulate at least 700 alternative splicing events in vivo, yet relatively little is known about the biologic consequences of NOVA action and in particular about functional differences between NOVA1 and NOVA2. Transcriptome-wide searches for isoform-specific functions, using NOVA1 and NOVA2 specific HITS-CLIP and RNA-seq data from mouse cortex lacking either NOVA isoform, reveals that NOVA2 uniquely regulates alternative splicing events of a series of axon guidance related genes during cortical development. Corresponding axonal pathfinding defects were specific to NOVA2 deficiency: Nova2-/- but not Nova1-/- mice had agenesis of the corpus callosum, and axonal outgrowth defects specific to ventral motoneuron axons and efferent innervation of the cochlea. Thus we have discovered that NOVA2 uniquely regulates alternative splicing of a coordinate set of transcripts encoding key components in cortical, brainstem and spinal axon guidance/outgrowth pathways during neural differentiation, with severe functional consequences in vivo.

No MeSH data available.


Related in: MedlinePlus