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Lhx5 controls mamillary differentiation in the developing hypothalamus of the mouse.

Heide M, Zhang Y, Zhou X, Zhao T, Miquelajáuregui A, Varela-Echavarría A, Alvarez-Bolado G - Front Neuroanat (2015)

Bottom Line: Microarray analysis and chromatin immunoprecipitation indicated that Lhx5 appears to be involved in Shh downregulation through Tbx3 and activates several MBO-specific regulator and effector genes.Finally, by tracing the caudal hypothalamic cell lineage we show that, in the Lhx5 mutant, at least some MBO cells are present but lack characteristic marker expression.Our work shows how the Lhx5 locus contributes to integrate regional specification pathways with downstream acquisition of neuronal identity in the MBO.

View Article: PubMed Central - PubMed

Affiliation: Institute of Anatomy and Cell Biology, University of Heidelberg Heidelberg, Germany.

ABSTRACT
Acquisition of specific neuronal identity by individual brain nuclei is a key step in brain development. However, how the mechanisms that confer neuronal identity are integrated with upstream regional specification networks is still mysterious. Expression of Sonic hedgehog (Shh), is required for hypothalamic specification and is later downregulated by Tbx3 to allow for the differentiation of the tubero-mamillary region. In this region, the mamillary body (MBO), is a large neuronal aggregate essential for memory formation. To clarify how MBO identity is acquired after regional specification, we investigated Lhx5, a transcription factor with restricted MBO expression. We first generated a hypomorph allele of Lhx5-in homozygotes, the MBO disappears after initial specification. Intriguingly, in these mutants, Tbx3 was downregulated and the Shh expression domain abnormally extended. Microarray analysis and chromatin immunoprecipitation indicated that Lhx5 appears to be involved in Shh downregulation through Tbx3 and activates several MBO-specific regulator and effector genes. Finally, by tracing the caudal hypothalamic cell lineage we show that, in the Lhx5 mutant, at least some MBO cells are present but lack characteristic marker expression. Our work shows how the Lhx5 locus contributes to integrate regional specification pathways with downstream acquisition of neuronal identity in the MBO.

No MeSH data available.


Tbx3 expression is reduced in the Lhx5fl∕fl mutant. (A–F)in situ detection of Tbx3 expression in Lhx5fl∕+ and Lhx5fl∕fl mutant E12.5 embryos. Three different medio-lateral levels are shown (from top to bottom). Arrows indicate the boundaries of Tbx3 expression in (A–D) and the lateral extension of the Tbx3 expression domain in (E,F). (G–N) Immunohistochemistry against LHX1/5 (red) and neurofilaments (green) in Tbx3 −∕− and control E14.5 embryos, counterstaining with DAPI. (G,J) show detection of LHX1/5 in the MBO of control (G) and Tbx3−∕− mutant (J). (H,K) show LHX1/5 plus neurofilaments in the MBO of control (H) and Tbx3−∕− mutant (K). (I,L) show the same panels as in (H,K) with thick lines indicating the MBO profile and thin lines indicating the principal mamillary tract axons. (M,N) show a higher magnification view of the axons of the principal mamillary tract in controls and mutants. Arrowheads indicate the axons of the principal mamillary tract. Asterisks indicate the position of the MBO. Scale bars: (A–F), 500 μm; (E–N), 100 μm.
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Figure 8: Tbx3 expression is reduced in the Lhx5fl∕fl mutant. (A–F)in situ detection of Tbx3 expression in Lhx5fl∕+ and Lhx5fl∕fl mutant E12.5 embryos. Three different medio-lateral levels are shown (from top to bottom). Arrows indicate the boundaries of Tbx3 expression in (A–D) and the lateral extension of the Tbx3 expression domain in (E,F). (G–N) Immunohistochemistry against LHX1/5 (red) and neurofilaments (green) in Tbx3 −∕− and control E14.5 embryos, counterstaining with DAPI. (G,J) show detection of LHX1/5 in the MBO of control (G) and Tbx3−∕− mutant (J). (H,K) show LHX1/5 plus neurofilaments in the MBO of control (H) and Tbx3−∕− mutant (K). (I,L) show the same panels as in (H,K) with thick lines indicating the MBO profile and thin lines indicating the principal mamillary tract axons. (M,N) show a higher magnification view of the axons of the principal mamillary tract in controls and mutants. Arrowheads indicate the axons of the principal mamillary tract. Asterisks indicate the position of the MBO. Scale bars: (A–F), 500 μm; (E–N), 100 μm.

Mentions: One of these genes, Tbx3, has an important role in the development of the hypothalamus (Manning et al., 2006; Trowe et al., 2013). We examined the expression of Tbx3 at three different medio-lateral levels in our mutants at E12.5 (Figures 8A–F) and found a strong reduction in the expression domain in the mutant. This reduction affected not only the rostro-caudal extension of the midline (Figures 8A,C vs. Figures 8B,D) but was also evident at more lateral levels (Figures 8E,F). We then hypothesized that Tbx3 is involved in MBO development, and on this basis predicted MBO defects in Tbx3-deficient brains. Examination of the hypothalamus of Tbx3 mutant mice (Hoogaars et al., 2007) at E14.5 showed a reduced MBO with abnormal morphology (Figures 8G–L) as well as a strong reduction in axonal projections (Figures 8M,N). Since the Tbx3 mutant embryos die before birth, usually around E14.5, we could not ascertain the possible total loss of the MBO at later stages.


Lhx5 controls mamillary differentiation in the developing hypothalamus of the mouse.

Heide M, Zhang Y, Zhou X, Zhao T, Miquelajáuregui A, Varela-Echavarría A, Alvarez-Bolado G - Front Neuroanat (2015)

Tbx3 expression is reduced in the Lhx5fl∕fl mutant. (A–F)in situ detection of Tbx3 expression in Lhx5fl∕+ and Lhx5fl∕fl mutant E12.5 embryos. Three different medio-lateral levels are shown (from top to bottom). Arrows indicate the boundaries of Tbx3 expression in (A–D) and the lateral extension of the Tbx3 expression domain in (E,F). (G–N) Immunohistochemistry against LHX1/5 (red) and neurofilaments (green) in Tbx3 −∕− and control E14.5 embryos, counterstaining with DAPI. (G,J) show detection of LHX1/5 in the MBO of control (G) and Tbx3−∕− mutant (J). (H,K) show LHX1/5 plus neurofilaments in the MBO of control (H) and Tbx3−∕− mutant (K). (I,L) show the same panels as in (H,K) with thick lines indicating the MBO profile and thin lines indicating the principal mamillary tract axons. (M,N) show a higher magnification view of the axons of the principal mamillary tract in controls and mutants. Arrowheads indicate the axons of the principal mamillary tract. Asterisks indicate the position of the MBO. Scale bars: (A–F), 500 μm; (E–N), 100 μm.
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Figure 8: Tbx3 expression is reduced in the Lhx5fl∕fl mutant. (A–F)in situ detection of Tbx3 expression in Lhx5fl∕+ and Lhx5fl∕fl mutant E12.5 embryos. Three different medio-lateral levels are shown (from top to bottom). Arrows indicate the boundaries of Tbx3 expression in (A–D) and the lateral extension of the Tbx3 expression domain in (E,F). (G–N) Immunohistochemistry against LHX1/5 (red) and neurofilaments (green) in Tbx3 −∕− and control E14.5 embryos, counterstaining with DAPI. (G,J) show detection of LHX1/5 in the MBO of control (G) and Tbx3−∕− mutant (J). (H,K) show LHX1/5 plus neurofilaments in the MBO of control (H) and Tbx3−∕− mutant (K). (I,L) show the same panels as in (H,K) with thick lines indicating the MBO profile and thin lines indicating the principal mamillary tract axons. (M,N) show a higher magnification view of the axons of the principal mamillary tract in controls and mutants. Arrowheads indicate the axons of the principal mamillary tract. Asterisks indicate the position of the MBO. Scale bars: (A–F), 500 μm; (E–N), 100 μm.
Mentions: One of these genes, Tbx3, has an important role in the development of the hypothalamus (Manning et al., 2006; Trowe et al., 2013). We examined the expression of Tbx3 at three different medio-lateral levels in our mutants at E12.5 (Figures 8A–F) and found a strong reduction in the expression domain in the mutant. This reduction affected not only the rostro-caudal extension of the midline (Figures 8A,C vs. Figures 8B,D) but was also evident at more lateral levels (Figures 8E,F). We then hypothesized that Tbx3 is involved in MBO development, and on this basis predicted MBO defects in Tbx3-deficient brains. Examination of the hypothalamus of Tbx3 mutant mice (Hoogaars et al., 2007) at E14.5 showed a reduced MBO with abnormal morphology (Figures 8G–L) as well as a strong reduction in axonal projections (Figures 8M,N). Since the Tbx3 mutant embryos die before birth, usually around E14.5, we could not ascertain the possible total loss of the MBO at later stages.

Bottom Line: Microarray analysis and chromatin immunoprecipitation indicated that Lhx5 appears to be involved in Shh downregulation through Tbx3 and activates several MBO-specific regulator and effector genes.Finally, by tracing the caudal hypothalamic cell lineage we show that, in the Lhx5 mutant, at least some MBO cells are present but lack characteristic marker expression.Our work shows how the Lhx5 locus contributes to integrate regional specification pathways with downstream acquisition of neuronal identity in the MBO.

View Article: PubMed Central - PubMed

Affiliation: Institute of Anatomy and Cell Biology, University of Heidelberg Heidelberg, Germany.

ABSTRACT
Acquisition of specific neuronal identity by individual brain nuclei is a key step in brain development. However, how the mechanisms that confer neuronal identity are integrated with upstream regional specification networks is still mysterious. Expression of Sonic hedgehog (Shh), is required for hypothalamic specification and is later downregulated by Tbx3 to allow for the differentiation of the tubero-mamillary region. In this region, the mamillary body (MBO), is a large neuronal aggregate essential for memory formation. To clarify how MBO identity is acquired after regional specification, we investigated Lhx5, a transcription factor with restricted MBO expression. We first generated a hypomorph allele of Lhx5-in homozygotes, the MBO disappears after initial specification. Intriguingly, in these mutants, Tbx3 was downregulated and the Shh expression domain abnormally extended. Microarray analysis and chromatin immunoprecipitation indicated that Lhx5 appears to be involved in Shh downregulation through Tbx3 and activates several MBO-specific regulator and effector genes. Finally, by tracing the caudal hypothalamic cell lineage we show that, in the Lhx5 mutant, at least some MBO cells are present but lack characteristic marker expression. Our work shows how the Lhx5 locus contributes to integrate regional specification pathways with downstream acquisition of neuronal identity in the MBO.

No MeSH data available.