Widespread Contribution of Gdf7 Lineage to Cerebellar Cell Types and Implications for Hedgehog-Driven Medulloblastoma Formation

The roof plate is a specialized embryonic midline tissue of the central nervous system that functions as a signaling center regulating dorsal neural patterning. In the developing hindbrain, roof plate cells express Gdf7 and previous genetic fate mapping studies showed that these cells contribute mostly to non-neural choroid plexus epithelium. We demonstrate here that constitutive activation of the Sonic hedgehog signaling pathway in the Gdf7 lineage invariably leads to medulloblastoma. Lineage tracing analysis reveals that Gdf7-lineage cells not only are a source of choroid plexus epithelial cells, but are also present in the cerebellar rhombic lip and contribute to a subset of cerebellar granule neuron precursors, the presumed cell-of-origin for Sonic hedgehog-driven medulloblastoma. We further show that Gdf7-lineage cells also contribute to multiple neuronal and glial cell types in the cerebellum, including glutamatergic granule neurons, unipolar brush cells, Purkinje neurons, GABAergic interneurons, Bergmann glial cells, and white matter astrocytes. These findings establish hindbrain roof plate as a novel source of diverse neural cell types in the cerebellum that is also susceptible to oncogenic transformation by deregulated Sonic hedgehog signaling

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A Grid portal is a user’s point of access to a Grid system [3,5,6,9]. It provides an environment where the user can access Grid resources and services, execute and monitor Grid applications, and collaborate with other users. A number of Grid portals exist or are currently under construction including the NEESgrid Portal [14], the Alliance Portal [22], the GENIUS Portal for the European Data Grid [19], and the IeSE (Integrated e

<i>Gdf7^Cre/;SmoM2</i> mice develop medulloblastoma with CGNP features.

<p>(A–B″′) In situ hybridization of wild-type and <i>Gdf7^Cre/+;SmoM2</i> mutants. The aberrant tissue foci of <i>Gdf7^Cre/+;SmoM2</i> cerebella display high level Shh signaling as determined by <i>Gli1</i> and <i>Ptch1</i> expression, and <i>Nmyc</i> and <i>Math1</i> expression. (C–H″′) <i>Gdf7^Cre/+;SmoM2</i> mice develop medulloblastoma consisting of cells of the CGNP fate. Tumors in <i>Gdf7^Cre/+;SmoM2</i> mice and adult <i>Patched1^LacZ/+;SmoM2</i> mice appear very similar. Abbreviations: ML, molecular layer. PCL, Purkinje cell layer. IGL, internal granular layer. T, tumor region. N, normal cerebellum.</p

Roof plate cells give rise to a distinct population of cerebellar vermal radial glia cells.

<p>(A) Whole mount X-gal staining for ß-galactosidase in <i>Gdf7^Cre/+;ROSA^lacZ</i> embryos shows that Gdf7 lineage is present in the hindbrain roof plate at E10. (B) At E14.5, in addition to hindbrain choroid plexus, Gdf7 lineage also contributes to the ventricular zone of cerebellar vermis (red arrows). (C–F) In situ hybridization at E14.5 showing <i>Gdf7</i> and its transcriptional target <i>Msx1</i> are expressed in the cerebellar vermis. (G–H′) Fate-mapping of the Gdf7-lineage in <i>Gdf7^Cre/+;ROSA^eYFP</i> cerebella indicates that Gdf7 is present at E14.5 in a distinct population of vermal radial glia cells as highlighted by coexpression of YFP and BLBP (G-G′) or Sox2 (H-H′) Arrows indicate colocalization. (I–K) In situ hybridization at P18 shows that the <i>Math1+ Gdf7^Cre/+;SmoM2</i> tumor tissue does not express <i>Gdf7</i> or <i>Msx1</i>. Abbreviations: N, normal cerebellar tissue. T, tumor tissue. CB, cerebellum. uRL, upper rhombic lip.</p

Shh pathway activation in Gdf7-lineage cells leads to cerebellar hyperplasia.

<p><i>Gdf7^Cre/+;SmoM2</i> gain-of-function mutant mice exhibit cerebellar defects. (A–B′) Hematoxylin-eosin staining of wild-type and <i>Gdf7^Cre/+;SmoM2</i> mutants. Prior to P10 histological sections of mutants are similar to control. (C–E′) <i>Gdf7^Cre/+;SmoM2</i> mutants over 14 days old develop ectopic foci of densely packed cells within the molecular layer of their cerebella. Higher magnification view of these foci reveals no discernible layer organization and resemblance to neoplastic lesions. Arrows in (D) indicate regions of hypercellularity. Boxed regions E and E″ are magnified and shown in the right adjacent panels. Arrows in D′ indicate nuclear molding. Arrowheads in D′ indicate apoptotic nuclei. (F-F″) The ectopic foci consist of cells of the Gdf7-lineage as indicated by their expression of SmoM2-YFP. (G-G″) Ectopic foci do not express differentiated neuronal marker NeuN. Abbreviations: EGL, external granular layer. ML, molecular layer. IGL, internal granular layer. T, tumor region. N, normal cerebellum.</p

A subset of Gdf7-lineage cells express neural stem cell markers.

<p>(A–B″′) Many cells within the tumor tissue of <i>Gdf7^Cre/+;SmoM2</i> mice coexpress neural stem cell marker Nestin (red) and glial marker GFAP (green). Arrows indicate co-localization. (C–E) Tumor cell lines can be invariably established from <i>Gdf7^Cre/+;SmoM2</i> cerebella. (F–G″) These cells highly express multiple neural stem cell markers Nestin and Sox2 and can undergo serial passages. (H–J) Here, one representative colony is shown when cultured for 3 and 6 days. (K–N) <i>Gdf7^Cre/+;SmoM2</i> cultured cells can differentiate into mature cerebellar cell types upon switching to culture media containing 10% FBS.</p