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The lineage contribution and role of Gbx2 in spinal cord development.

Luu B, Ellisor D, Zervas M - PLoS ONE (2011)

Bottom Line: Using lineage tracing and molecular markers to follow Gbx2-mutant cells, we show that the loss of Gbx2 globally affects spinal cord patterning including the organization of interneuron progenitors.Finally, long-term lineage analysis reveals that the presence and timing of Gbx2 expression in interneuron progenitors results in the differential contribution to subtypes of terminally differentiated interneurons in the adult spinal cord.In a broader context, this study provides a direct link between spinal cord progenitors undergoing dynamic changes in molecular identity and terminal neuronal fate.

View Article: PubMed Central - PubMed

Affiliation: Division of Biology and Medicine, Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, Rhode Island, United States of America.

ABSTRACT

Background: Forging a relationship between progenitors with dynamically changing gene expression and their terminal fate is instructive for understanding the logic of how cell-type diversity is established. The mouse spinal cord is an ideal system to study these mechanisms in the context of developmental genetics and nervous system development. Here we focus on the Gastrulation homeobox 2 (Gbx2) transcription factor, which has not been explored in spinal cord development.

Methodology/principal findings: We determined the molecular identity of Gbx2-expressing spinal cord progenitors. We also utilized genetic inducible fate mapping to mark the Gbx2 lineage at different embryonic stages in vivo in mouse. Collectively, we uncover cell behaviors, cytoarchitectonic organization, and the terminal cell fate of the Gbx2 lineage. Notably, both ventral motor neurons and interneurons are derived from the Gbx2 lineage, but only during a short developmental period. Short-term fate mapping during mouse spinal cord development shows that Gbx2 expression is transient and is extinguished ventrally in a rostral to caudal gradient. Concomitantly, a permanent lineage restriction boundary ensures that spinal cord neurons derived from the Gbx2 lineage are confined to a dorsal compartment that is maintained in the adult and that this lineage generates inhibitory interneurons of the spinal cord. Using lineage tracing and molecular markers to follow Gbx2-mutant cells, we show that the loss of Gbx2 globally affects spinal cord patterning including the organization of interneuron progenitors. Finally, long-term lineage analysis reveals that the presence and timing of Gbx2 expression in interneuron progenitors results in the differential contribution to subtypes of terminally differentiated interneurons in the adult spinal cord.

Conclusions/significance: We illustrate the complex cellular nature of Gbx2 expression and lineage contribution to the mouse spinal cord. In a broader context, this study provides a direct link between spinal cord progenitors undergoing dynamic changes in molecular identity and terminal neuronal fate.

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Related in: MedlinePlus

Spatial distribution of the Gbx2 lineage in E12.5 spinal cord.(A–D) Gbx2-derived cells marked at E8.5 (ß-gal+, red) on sagittal sections of E12.5 spinal cord at the indicated levels. (E–G) Transverse sections of E12.5 spinal cord at the indicated levels showing ß-gal+ cells (red) that were marked at E9.5. (H–J) The Gbx2 lineage (ß-gal+, red) marked at E10.5 was confined to the dorsal spinal cord at all axial levels at E12.5. (K–M) Transverse sections showing Gbx2(GFP)+ cells at indicated levels in E12.5 spinal cord. (N–P) Comparison of the Gbx2 lineage (ß-gal+, red) marked at E8.5 (N), E9.5 (O), and E10.5 (P) versus Gbx2 expression (GFP+, green) in E12.5 spinal cord. (N) Four D-V Gbx2-derived sub-populations can be classified by the presence or absence of Gbx2: zones 1 and 3 are Gbx2-derived cells that persisted in Gbx2 expression while zones 2 and 4 have down-regulated Gbx2. (O) Gbx2-derived cells marked at E9.5 continued to express Gbx2(GFP) in dorsal spinal cord at E12.5 in contrast to few ventral cells. (P) The majority of Gbx2(GFP)-expressing cells marked at E10.5 were confined to a dorsal domain and continued to express Gbx2(GFP).
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pone-0020940-g002: Spatial distribution of the Gbx2 lineage in E12.5 spinal cord.(A–D) Gbx2-derived cells marked at E8.5 (ß-gal+, red) on sagittal sections of E12.5 spinal cord at the indicated levels. (E–G) Transverse sections of E12.5 spinal cord at the indicated levels showing ß-gal+ cells (red) that were marked at E9.5. (H–J) The Gbx2 lineage (ß-gal+, red) marked at E10.5 was confined to the dorsal spinal cord at all axial levels at E12.5. (K–M) Transverse sections showing Gbx2(GFP)+ cells at indicated levels in E12.5 spinal cord. (N–P) Comparison of the Gbx2 lineage (ß-gal+, red) marked at E8.5 (N), E9.5 (O), and E10.5 (P) versus Gbx2 expression (GFP+, green) in E12.5 spinal cord. (N) Four D-V Gbx2-derived sub-populations can be classified by the presence or absence of Gbx2: zones 1 and 3 are Gbx2-derived cells that persisted in Gbx2 expression while zones 2 and 4 have down-regulated Gbx2. (O) Gbx2-derived cells marked at E9.5 continued to express Gbx2(GFP) in dorsal spinal cord at E12.5 in contrast to few ventral cells. (P) The majority of Gbx2(GFP)-expressing cells marked at E10.5 were confined to a dorsal domain and continued to express Gbx2(GFP).

Mentions: Gbx2(GFP) was strongly expressed along the A-P axis of the E12.5 spinal cord in a broad dorsal domain and in a ventral strip on sagittal sections (Figure 1T–V). The Gbx2(GFP) expression domain on transverse sections at E12.5 spanned the entire medial-lateral axis in the dorsal spinal cord (Figures 1W,X and 2K–M). In contrast, Gbx2(GFP) expression ventrally was limited to two well-delineated bi-lateral columns (Figures 1W,X and 2K–M). Notably, the Gbx2(GFP)+ domains exactly recapitulated the RNA in situ pattern of Gbx2 expression at E12.5 (Figure S1). Compared to the expression pattern at E10.5, dorsal Gbx2(GFP) expression at E12.5 spanned a proportionally broader D-V domain and extended up to the dorsal-most point of spinal cord, excluding the roof plate (Figures 1K–R,1W–X). The Gbx2(GFP) cells dorsally expressed Pax2, a class-A transcription factor that defines late-born inhibitory interneurons at E12.5 [32] (Figure 1W,W′). Gbx2(GFP) expression co-localized more extensively with Pax2 dorsolaterally indicating that Gbx2(GFP) was also expressed in differentiating earlier-born inhibitory interneurons that have settled in the mantle and marginal zones (Figure 1W,W′). The most ventral Gbx2(GFP)-expressing cells only rarely co-expressed Pax2 (Figure 1W″). Gbx2(GFP) at E12.5 expression did not co-localize with dorsal Isl1/2 (Figure 1X,X′), which is a marker for dorsal excitatory interneurons [30]. To assess Gbx2(GFP) expression in motor neuron populations we analyzed ventral Isl1/2, which is a marker for all differentiating motor neurons in the ventral spinal cord [33]. Ventral-lateral Gbx2(GFP) expressing cells were nested within the Isl1/2+ medial and lateral motor columns (Figure 1X,X″), but did not did not co-localize with Isl1/2. Therefore, Gbx2(GFP) expression does not identify motor neuron sub-populations at E12.5, but Gbx2(GFP) expression at E12.5 defines early and late-born dorsal inhibitory interneuron sub-populations.


The lineage contribution and role of Gbx2 in spinal cord development.

Luu B, Ellisor D, Zervas M - PLoS ONE (2011)

Spatial distribution of the Gbx2 lineage in E12.5 spinal cord.(A–D) Gbx2-derived cells marked at E8.5 (ß-gal+, red) on sagittal sections of E12.5 spinal cord at the indicated levels. (E–G) Transverse sections of E12.5 spinal cord at the indicated levels showing ß-gal+ cells (red) that were marked at E9.5. (H–J) The Gbx2 lineage (ß-gal+, red) marked at E10.5 was confined to the dorsal spinal cord at all axial levels at E12.5. (K–M) Transverse sections showing Gbx2(GFP)+ cells at indicated levels in E12.5 spinal cord. (N–P) Comparison of the Gbx2 lineage (ß-gal+, red) marked at E8.5 (N), E9.5 (O), and E10.5 (P) versus Gbx2 expression (GFP+, green) in E12.5 spinal cord. (N) Four D-V Gbx2-derived sub-populations can be classified by the presence or absence of Gbx2: zones 1 and 3 are Gbx2-derived cells that persisted in Gbx2 expression while zones 2 and 4 have down-regulated Gbx2. (O) Gbx2-derived cells marked at E9.5 continued to express Gbx2(GFP) in dorsal spinal cord at E12.5 in contrast to few ventral cells. (P) The majority of Gbx2(GFP)-expressing cells marked at E10.5 were confined to a dorsal domain and continued to express Gbx2(GFP).
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Related In: Results  -  Collection

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pone-0020940-g002: Spatial distribution of the Gbx2 lineage in E12.5 spinal cord.(A–D) Gbx2-derived cells marked at E8.5 (ß-gal+, red) on sagittal sections of E12.5 spinal cord at the indicated levels. (E–G) Transverse sections of E12.5 spinal cord at the indicated levels showing ß-gal+ cells (red) that were marked at E9.5. (H–J) The Gbx2 lineage (ß-gal+, red) marked at E10.5 was confined to the dorsal spinal cord at all axial levels at E12.5. (K–M) Transverse sections showing Gbx2(GFP)+ cells at indicated levels in E12.5 spinal cord. (N–P) Comparison of the Gbx2 lineage (ß-gal+, red) marked at E8.5 (N), E9.5 (O), and E10.5 (P) versus Gbx2 expression (GFP+, green) in E12.5 spinal cord. (N) Four D-V Gbx2-derived sub-populations can be classified by the presence or absence of Gbx2: zones 1 and 3 are Gbx2-derived cells that persisted in Gbx2 expression while zones 2 and 4 have down-regulated Gbx2. (O) Gbx2-derived cells marked at E9.5 continued to express Gbx2(GFP) in dorsal spinal cord at E12.5 in contrast to few ventral cells. (P) The majority of Gbx2(GFP)-expressing cells marked at E10.5 were confined to a dorsal domain and continued to express Gbx2(GFP).
Mentions: Gbx2(GFP) was strongly expressed along the A-P axis of the E12.5 spinal cord in a broad dorsal domain and in a ventral strip on sagittal sections (Figure 1T–V). The Gbx2(GFP) expression domain on transverse sections at E12.5 spanned the entire medial-lateral axis in the dorsal spinal cord (Figures 1W,X and 2K–M). In contrast, Gbx2(GFP) expression ventrally was limited to two well-delineated bi-lateral columns (Figures 1W,X and 2K–M). Notably, the Gbx2(GFP)+ domains exactly recapitulated the RNA in situ pattern of Gbx2 expression at E12.5 (Figure S1). Compared to the expression pattern at E10.5, dorsal Gbx2(GFP) expression at E12.5 spanned a proportionally broader D-V domain and extended up to the dorsal-most point of spinal cord, excluding the roof plate (Figures 1K–R,1W–X). The Gbx2(GFP) cells dorsally expressed Pax2, a class-A transcription factor that defines late-born inhibitory interneurons at E12.5 [32] (Figure 1W,W′). Gbx2(GFP) expression co-localized more extensively with Pax2 dorsolaterally indicating that Gbx2(GFP) was also expressed in differentiating earlier-born inhibitory interneurons that have settled in the mantle and marginal zones (Figure 1W,W′). The most ventral Gbx2(GFP)-expressing cells only rarely co-expressed Pax2 (Figure 1W″). Gbx2(GFP) at E12.5 expression did not co-localize with dorsal Isl1/2 (Figure 1X,X′), which is a marker for dorsal excitatory interneurons [30]. To assess Gbx2(GFP) expression in motor neuron populations we analyzed ventral Isl1/2, which is a marker for all differentiating motor neurons in the ventral spinal cord [33]. Ventral-lateral Gbx2(GFP) expressing cells were nested within the Isl1/2+ medial and lateral motor columns (Figure 1X,X″), but did not did not co-localize with Isl1/2. Therefore, Gbx2(GFP) expression does not identify motor neuron sub-populations at E12.5, but Gbx2(GFP) expression at E12.5 defines early and late-born dorsal inhibitory interneuron sub-populations.

Bottom Line: Using lineage tracing and molecular markers to follow Gbx2-mutant cells, we show that the loss of Gbx2 globally affects spinal cord patterning including the organization of interneuron progenitors.Finally, long-term lineage analysis reveals that the presence and timing of Gbx2 expression in interneuron progenitors results in the differential contribution to subtypes of terminally differentiated interneurons in the adult spinal cord.In a broader context, this study provides a direct link between spinal cord progenitors undergoing dynamic changes in molecular identity and terminal neuronal fate.

View Article: PubMed Central - PubMed

Affiliation: Division of Biology and Medicine, Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, Rhode Island, United States of America.

ABSTRACT

Background: Forging a relationship between progenitors with dynamically changing gene expression and their terminal fate is instructive for understanding the logic of how cell-type diversity is established. The mouse spinal cord is an ideal system to study these mechanisms in the context of developmental genetics and nervous system development. Here we focus on the Gastrulation homeobox 2 (Gbx2) transcription factor, which has not been explored in spinal cord development.

Methodology/principal findings: We determined the molecular identity of Gbx2-expressing spinal cord progenitors. We also utilized genetic inducible fate mapping to mark the Gbx2 lineage at different embryonic stages in vivo in mouse. Collectively, we uncover cell behaviors, cytoarchitectonic organization, and the terminal cell fate of the Gbx2 lineage. Notably, both ventral motor neurons and interneurons are derived from the Gbx2 lineage, but only during a short developmental period. Short-term fate mapping during mouse spinal cord development shows that Gbx2 expression is transient and is extinguished ventrally in a rostral to caudal gradient. Concomitantly, a permanent lineage restriction boundary ensures that spinal cord neurons derived from the Gbx2 lineage are confined to a dorsal compartment that is maintained in the adult and that this lineage generates inhibitory interneurons of the spinal cord. Using lineage tracing and molecular markers to follow Gbx2-mutant cells, we show that the loss of Gbx2 globally affects spinal cord patterning including the organization of interneuron progenitors. Finally, long-term lineage analysis reveals that the presence and timing of Gbx2 expression in interneuron progenitors results in the differential contribution to subtypes of terminally differentiated interneurons in the adult spinal cord.

Conclusions/significance: We illustrate the complex cellular nature of Gbx2 expression and lineage contribution to the mouse spinal cord. In a broader context, this study provides a direct link between spinal cord progenitors undergoing dynamic changes in molecular identity and terminal neuronal fate.

Show MeSH
Related in: MedlinePlus