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Cadherin 2/4 signaling via PTP1B and catenins is crucial for nucleokinesis during radial neuronal migration in the neocortex.

Martinez-Garay I, Gil-Sanz C, Franco SJ, Espinosa A, Molnár Z, Mueller U - Development (2016)

Bottom Line: Surprisingly, perturbation of cadherin-mediated signaling does not affect the formation and extension of leading processes of migrating neocortical neurons.Instead, movement of the cell body and nucleus (nucleokinesis) is disrupted.Taken together, our findings indicate that cadherin-mediated signaling to the cytoskeleton is crucial for nucleokinesis of neocortical projection neurons during their radial migration.

View Article: PubMed Central - PubMed

Affiliation: Molecular and Cellular Neuroscience Department, Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, CA 92037, USA martinezgarayi@cardiff.ac.uk umueller@scripps.edu.

No MeSH data available.


Related in: MedlinePlus

Functional interactions between CDH2 and LIS1. (A) Stack of confocal images of neurons expressing CDH2 (green), LIS1 (red) and BFP. EGFP-tagged CDH2 was co-electroporated with HA-tagged LIS1 and BFP at E14.5. CDH2 was visualized at E17.5 by EGFP fluorescence and LIS1 by staining with HA antibodies. Panels on the right are single confocal sections of the areas marked as A′ and A″. CDH2-EGFP and HA-LIS1 colocalize in the leading process (A′) and cell soma (A″). Arrows point to the leading processes of several neurons. (B) LIS1 localization is altered in neurons expressing DN-CDH. HA-tagged LIS1 was co-electroporated with either a control plasmid or DN-CDH at E14.5 and brains were analyzed 3 days later. DN-CDH-expressing neurons show increased HA-LIS1 staining in the leading process. B′ and B″ panels are single and combined channel images of the boxed areas in the main image. Dotted lines indicate leading processes. (C) Quantification of the relative HA (red) average fluorescence intensity in the leading processes versus soma of HA-LIS1 and control or DN-CDH co-electroporated neurons. *P<0.001 by Student's t-test. Fluorescence intensity was measured in 53 (control, four different brains) and 59 (DN-CDH, four different brains) neurons. (D) Co-expression of DCX-Lis1 partially rescues the migration defect caused by expression of DN-CDH. DN-CDH was co-electroporated either with a control plasmid or with DCX-Lis1-i-EGFP at E14.5 and the position of the electroporated cells was assessed at E18.5. Electroporated neurons are shown in black. (E) Quantification (mean±s.e.m.) of the data in D. *P<0.01 by Student's t-test. Neurons were counted in three brain slices from each of four animals obtained from three independent electroporation experiments. (F) High magnifications of the neurons expressing DN-CDH+control plasmid or DN-CDH+LIS1. Note the difference in the length of the leading processes between the two conditions. (G) Quantification of process length of the neurons in F. *P<0.01 by Student's t-test. Process length was measured in 44 (DN-CDH+EGFP, four different brains) and 48 (DN-CDH+DCX-Lis1, five different brains) neurons. MZ, marginal zone. Scale bars: 20 μm (A,B); 10 μm (A′,A″); 100 μm (D); 50 μm (F).
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DEV132456F8: Functional interactions between CDH2 and LIS1. (A) Stack of confocal images of neurons expressing CDH2 (green), LIS1 (red) and BFP. EGFP-tagged CDH2 was co-electroporated with HA-tagged LIS1 and BFP at E14.5. CDH2 was visualized at E17.5 by EGFP fluorescence and LIS1 by staining with HA antibodies. Panels on the right are single confocal sections of the areas marked as A′ and A″. CDH2-EGFP and HA-LIS1 colocalize in the leading process (A′) and cell soma (A″). Arrows point to the leading processes of several neurons. (B) LIS1 localization is altered in neurons expressing DN-CDH. HA-tagged LIS1 was co-electroporated with either a control plasmid or DN-CDH at E14.5 and brains were analyzed 3 days later. DN-CDH-expressing neurons show increased HA-LIS1 staining in the leading process. B′ and B″ panels are single and combined channel images of the boxed areas in the main image. Dotted lines indicate leading processes. (C) Quantification of the relative HA (red) average fluorescence intensity in the leading processes versus soma of HA-LIS1 and control or DN-CDH co-electroporated neurons. *P<0.001 by Student's t-test. Fluorescence intensity was measured in 53 (control, four different brains) and 59 (DN-CDH, four different brains) neurons. (D) Co-expression of DCX-Lis1 partially rescues the migration defect caused by expression of DN-CDH. DN-CDH was co-electroporated either with a control plasmid or with DCX-Lis1-i-EGFP at E14.5 and the position of the electroporated cells was assessed at E18.5. Electroporated neurons are shown in black. (E) Quantification (mean±s.e.m.) of the data in D. *P<0.01 by Student's t-test. Neurons were counted in three brain slices from each of four animals obtained from three independent electroporation experiments. (F) High magnifications of the neurons expressing DN-CDH+control plasmid or DN-CDH+LIS1. Note the difference in the length of the leading processes between the two conditions. (G) Quantification of process length of the neurons in F. *P<0.01 by Student's t-test. Process length was measured in 44 (DN-CDH+EGFP, four different brains) and 48 (DN-CDH+DCX-Lis1, five different brains) neurons. MZ, marginal zone. Scale bars: 20 μm (A,B); 10 μm (A′,A″); 100 μm (D); 50 μm (F).

Mentions: Knockdown of LIS1 (also known as PAFAH1B1) leads to a similar defect in radial migration as reported here, in that the LIS1-deficient neurons form elongated leading processes but fail to translocate their nucleus along these processes (Tsai et al., 2005; Youn et al., 2009). LIS1 is thought to act in concert with its binding partner dynein 1 to regulate migration by effects on microtubules, which are targeted towards sites of cell-cell adhesion where dynein interacts with β-catenin (Ligon et al., 2001). Since proper organization of the microtubule cytoskeleton is crucial for nuclear migration, we hypothesized that defects in cadherin signaling might also affect microtubules, where the dynein-LIS1 complex might provide a link between microtubules and cadherins. To test this model, we first determined whether LIS1 and CDH2 colocalize in migrating neurons. Since no suitable antibodies for LIS1 were available for high-resolution immunolocalization studies, we generated expression vectors for a GFP-tagged version of CDH2 and a HA-tagged version of LIS1. We co-electroporated the constructs into migrating neurons at E14.5, together with BFP to delineate the neuronal cell bodies and processes (Fig. 8A). We then used EGFP fluorescence and antibodies to HA to analyze the distribution of CDH2-EGFP and HA-LIS1 at E17.5 during the active migration phase of the electroporated neurons. CDH2-GFP and HA-LIS1 colocalized in the cell bodies and the leading processes of radially migrating neurons (Fig. 8A). Next, we determined whether expression of DN-CDH did alter LIS1 localization in migrating neurons. DN-CDH-expressing cells showed significantly more LIS1 in their leading processes compared with control neurons (Fig. 8B,C). In addition, neurons overexpressing LIS1, although still able to migrate into the cortical plate, displayed altered leading processes that appeared thinner and shorter than those of control cells. However, neurons that had been electroporated with DN-CDH showed no such alterations in their leading process when they overexpressed LIS1 (Fig. S9). Strikingly, when we co-expressed DCX-LIS1 with DN-CDH, the migratory defect caused by expression of DN-CDH was significantly but not completely rescued (Fig. 8D,E). Our electroporation of DCX-LIS1 alone did not change the percentage of neurons reaching the CP after 4 days (Fig. S9), suggesting that CDH2 and LIS1 act in a common pathway to regulate radial migration. In fact, expression of DCX-LIS1 not only increases the number of DN-CDH electroporated neurons in the CP 4 days after electroporation, but it also decreases the length of their leading processes, bringing their length closer to that of control cells (Fig. 8F,G; compare with control cell length in Fig. 5E).Fig. 8.


Cadherin 2/4 signaling via PTP1B and catenins is crucial for nucleokinesis during radial neuronal migration in the neocortex.

Martinez-Garay I, Gil-Sanz C, Franco SJ, Espinosa A, Molnár Z, Mueller U - Development (2016)

Functional interactions between CDH2 and LIS1. (A) Stack of confocal images of neurons expressing CDH2 (green), LIS1 (red) and BFP. EGFP-tagged CDH2 was co-electroporated with HA-tagged LIS1 and BFP at E14.5. CDH2 was visualized at E17.5 by EGFP fluorescence and LIS1 by staining with HA antibodies. Panels on the right are single confocal sections of the areas marked as A′ and A″. CDH2-EGFP and HA-LIS1 colocalize in the leading process (A′) and cell soma (A″). Arrows point to the leading processes of several neurons. (B) LIS1 localization is altered in neurons expressing DN-CDH. HA-tagged LIS1 was co-electroporated with either a control plasmid or DN-CDH at E14.5 and brains were analyzed 3 days later. DN-CDH-expressing neurons show increased HA-LIS1 staining in the leading process. B′ and B″ panels are single and combined channel images of the boxed areas in the main image. Dotted lines indicate leading processes. (C) Quantification of the relative HA (red) average fluorescence intensity in the leading processes versus soma of HA-LIS1 and control or DN-CDH co-electroporated neurons. *P<0.001 by Student's t-test. Fluorescence intensity was measured in 53 (control, four different brains) and 59 (DN-CDH, four different brains) neurons. (D) Co-expression of DCX-Lis1 partially rescues the migration defect caused by expression of DN-CDH. DN-CDH was co-electroporated either with a control plasmid or with DCX-Lis1-i-EGFP at E14.5 and the position of the electroporated cells was assessed at E18.5. Electroporated neurons are shown in black. (E) Quantification (mean±s.e.m.) of the data in D. *P<0.01 by Student's t-test. Neurons were counted in three brain slices from each of four animals obtained from three independent electroporation experiments. (F) High magnifications of the neurons expressing DN-CDH+control plasmid or DN-CDH+LIS1. Note the difference in the length of the leading processes between the two conditions. (G) Quantification of process length of the neurons in F. *P<0.01 by Student's t-test. Process length was measured in 44 (DN-CDH+EGFP, four different brains) and 48 (DN-CDH+DCX-Lis1, five different brains) neurons. MZ, marginal zone. Scale bars: 20 μm (A,B); 10 μm (A′,A″); 100 μm (D); 50 μm (F).
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DEV132456F8: Functional interactions between CDH2 and LIS1. (A) Stack of confocal images of neurons expressing CDH2 (green), LIS1 (red) and BFP. EGFP-tagged CDH2 was co-electroporated with HA-tagged LIS1 and BFP at E14.5. CDH2 was visualized at E17.5 by EGFP fluorescence and LIS1 by staining with HA antibodies. Panels on the right are single confocal sections of the areas marked as A′ and A″. CDH2-EGFP and HA-LIS1 colocalize in the leading process (A′) and cell soma (A″). Arrows point to the leading processes of several neurons. (B) LIS1 localization is altered in neurons expressing DN-CDH. HA-tagged LIS1 was co-electroporated with either a control plasmid or DN-CDH at E14.5 and brains were analyzed 3 days later. DN-CDH-expressing neurons show increased HA-LIS1 staining in the leading process. B′ and B″ panels are single and combined channel images of the boxed areas in the main image. Dotted lines indicate leading processes. (C) Quantification of the relative HA (red) average fluorescence intensity in the leading processes versus soma of HA-LIS1 and control or DN-CDH co-electroporated neurons. *P<0.001 by Student's t-test. Fluorescence intensity was measured in 53 (control, four different brains) and 59 (DN-CDH, four different brains) neurons. (D) Co-expression of DCX-Lis1 partially rescues the migration defect caused by expression of DN-CDH. DN-CDH was co-electroporated either with a control plasmid or with DCX-Lis1-i-EGFP at E14.5 and the position of the electroporated cells was assessed at E18.5. Electroporated neurons are shown in black. (E) Quantification (mean±s.e.m.) of the data in D. *P<0.01 by Student's t-test. Neurons were counted in three brain slices from each of four animals obtained from three independent electroporation experiments. (F) High magnifications of the neurons expressing DN-CDH+control plasmid or DN-CDH+LIS1. Note the difference in the length of the leading processes between the two conditions. (G) Quantification of process length of the neurons in F. *P<0.01 by Student's t-test. Process length was measured in 44 (DN-CDH+EGFP, four different brains) and 48 (DN-CDH+DCX-Lis1, five different brains) neurons. MZ, marginal zone. Scale bars: 20 μm (A,B); 10 μm (A′,A″); 100 μm (D); 50 μm (F).
Mentions: Knockdown of LIS1 (also known as PAFAH1B1) leads to a similar defect in radial migration as reported here, in that the LIS1-deficient neurons form elongated leading processes but fail to translocate their nucleus along these processes (Tsai et al., 2005; Youn et al., 2009). LIS1 is thought to act in concert with its binding partner dynein 1 to regulate migration by effects on microtubules, which are targeted towards sites of cell-cell adhesion where dynein interacts with β-catenin (Ligon et al., 2001). Since proper organization of the microtubule cytoskeleton is crucial for nuclear migration, we hypothesized that defects in cadherin signaling might also affect microtubules, where the dynein-LIS1 complex might provide a link between microtubules and cadherins. To test this model, we first determined whether LIS1 and CDH2 colocalize in migrating neurons. Since no suitable antibodies for LIS1 were available for high-resolution immunolocalization studies, we generated expression vectors for a GFP-tagged version of CDH2 and a HA-tagged version of LIS1. We co-electroporated the constructs into migrating neurons at E14.5, together with BFP to delineate the neuronal cell bodies and processes (Fig. 8A). We then used EGFP fluorescence and antibodies to HA to analyze the distribution of CDH2-EGFP and HA-LIS1 at E17.5 during the active migration phase of the electroporated neurons. CDH2-GFP and HA-LIS1 colocalized in the cell bodies and the leading processes of radially migrating neurons (Fig. 8A). Next, we determined whether expression of DN-CDH did alter LIS1 localization in migrating neurons. DN-CDH-expressing cells showed significantly more LIS1 in their leading processes compared with control neurons (Fig. 8B,C). In addition, neurons overexpressing LIS1, although still able to migrate into the cortical plate, displayed altered leading processes that appeared thinner and shorter than those of control cells. However, neurons that had been electroporated with DN-CDH showed no such alterations in their leading process when they overexpressed LIS1 (Fig. S9). Strikingly, when we co-expressed DCX-LIS1 with DN-CDH, the migratory defect caused by expression of DN-CDH was significantly but not completely rescued (Fig. 8D,E). Our electroporation of DCX-LIS1 alone did not change the percentage of neurons reaching the CP after 4 days (Fig. S9), suggesting that CDH2 and LIS1 act in a common pathway to regulate radial migration. In fact, expression of DCX-LIS1 not only increases the number of DN-CDH electroporated neurons in the CP 4 days after electroporation, but it also decreases the length of their leading processes, bringing their length closer to that of control cells (Fig. 8F,G; compare with control cell length in Fig. 5E).Fig. 8.

Bottom Line: Surprisingly, perturbation of cadherin-mediated signaling does not affect the formation and extension of leading processes of migrating neocortical neurons.Instead, movement of the cell body and nucleus (nucleokinesis) is disrupted.Taken together, our findings indicate that cadherin-mediated signaling to the cytoskeleton is crucial for nucleokinesis of neocortical projection neurons during their radial migration.

View Article: PubMed Central - PubMed

Affiliation: Molecular and Cellular Neuroscience Department, Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, CA 92037, USA martinezgarayi@cardiff.ac.uk umueller@scripps.edu.

No MeSH data available.


Related in: MedlinePlus