Neural stem cells, which exhibit self-renewal and multipotentiality, are generated in early embryonic brains and maintained throughout the lifespan. The mechanisms of their generation and maintenance are largely unknown. Here, we show that neural stem cells are generated independent of RBP-J , a key molecule in Notch signaling, by using RBP-J −/− embryonic stem cells in an embryonic stem cell-derived neurosphere assay. However, Notch pathway molecules are essential for the maintenance of neural stem cells; they are depleted in the early embryonic brains of RBP-J −/− or Notch1 −/− mice. Neural stem cells also are depleted in embryonic brains deficient for the presenilin1 (PS1) gene, a key regulator in Notch signaling, and are reduced in PS1 +/− adult brains. Both neuronal and glial differentiation in vitro were enhanced by attenuation of Notch signaling and suppressed by expressing an active form of Notch1. These data are consistent with a role for Notch signaling in the maintenance of the neural stem cell, and inconsistent with a role in a neuronal/glial fate switch.[Key Words: Presenilin; RBP-J ; embryonic stem cell; self-renewal; multipotentiality; cell cycle time]Received August 31, 2001; revised version accepted February 11, 2002. Neural stem cells, which are considered the ultimate lineage precursors to all neuronal and glial cells in the mammalian nervous system, are present not only in the developing brain but also in the adult brain Gage 2000). Although neural stem cells have a fundamental role in generating cellular diversity in the developing mammalian nervous system and in maintaining normal brain functions in adult brains (Lois and Alvarez-Buylla 1994;Tropepe et al. 1999;Shors et al. 2001), little is known concerning molecular mechanisms regulating the generation and maintenance of neural stem cells. In vitro, single neural stem cells proliferate to form clonally derived floating sphere colonies (neurospheres), which contain cells that, upon dissociation into single cells, give rise to new sphere colonies (self-renewal) and cells that can differentiate into neurons or glia (multipotentiality). Fibroblast growth factor-2 (FGF2)-responsive neural stem cells first appear in vivo at embryonic day (E) 8.5 and a separate and additive population of epidermal growth factor (EGF)-responsive neural stem cells arises from the earlier born FGF2-responsive stem cells by asymmetric division between E11 and E13 (Burrows et al. 1997;Mayer-Proschel et al. 1997;Tropepe et al. 1999). Both FGF2-responsive and EGF-responsive neural stem cells expand their populations and extend their cell cycle times during later embryogenesis (Martens et al. 2000). In the adult forebrain, neural stem cells are present as a relatively quiescent subpopulation in the subependyma, a remnant of the embryonic germinal zone (Morshead et al. 1994). This population persists into senescence, and the number is maintained throughout life (Tropepe et al. 1997). Thus, the generation and the size of the neural stem-cell population are tightly regul...

The human skeleton is affected by mutations in Low-density lipoprotein Receptor-related Protein 5 (LRP5). To understand how LRP5 influences bone properties, we generated mice with inducible Lrp5 mutations that cause high bone mass and low bone mass phenotypes in humans. We conditionally-induced Lrp5 mutations in osteocytes and found that bone properties in these mice were comparable to bone properties in mice with inherited mutations. We also conditionally-induced an Lrp5 mutation in cells that contribute to the appendicular skeleton, and not to the axial skeleton, and we observed bone properties were altered in the limbs, and not in the spine. These data indicate that Lrp5 signaling functions locally and suggest increasing LRP5 signaling in mature bone cells as a strategy to treat human low bone mass disorders, such as osteoporosis.

Dorsal dermis and epaxial muscle have been shown to arise from the central dermomyotome in the chick. En1 is a homeobox transcription factor gene expressed in the central dermomyotome. We show by genetic fate mapping in the mouse that En1-expressing cells of the central dermomyotome give rise to dorsal dermis and epaxial muscle and, unexpectedly, to interscapular brown fat. Thus, the En1-expressing central dermomyotome normally gives rise to three distinct fates in mice. Wnt signals are important in early stages of dermomyotome development, but the signal that acts to specify the dermal fate has not been identified. Using a reporter transgene for Wnt signal transduction, we show that the En1-expressing cells directly underneath the surface ectoderm transduce Wnt signals. When the essential Wnt transducer beta-catenin is mutated in En1 cells, it results in the loss of Dermo1-expressing dorsal dermal progenitors and dermis. Conversely, when beta-catenin was activated in En1 cells, it induces Dermo1 expression in all cells of the En1 domain and disrupts muscle gene expression. Our results indicate that the mouse central dermomyotome gives rise to dermis, muscle, and brown fat, and that Wnt signalling normally instructs cells to select the dorsal dermal fate.

Cajal bodies (CBs) are nuclear suborganelles involved in the biogenesis of small nuclear ribonucleoproteins (snRNPs). In addition to snRNPs, they are highly enriched in basal transcription and cell cycle factors, the nucleolar proteins fibrillarin (Fb) and Nopp140 (Nopp), the survival motor neuron (SMN) protein complex, and the CB marker protein, p80 coilin. We report the generation of knockout mice lacking the COOH-terminal 487 amino acids of coilin. Northern and Western blot analyses demonstrate that we have successfully removed the full-length coilin protein from the knockout animals. Some homozygous mutant animals are viable, but their numbers are reduced significantly when crossed to inbred backgrounds. Analysis of tissues and cell lines from mutant animals reveals the presence of extranucleolar foci that contain Fb and Nopp but not other typical nucleolar markers. These so-called “residual” CBs neither condense Sm proteins nor recruit members of the SMN protein complex. Transient expression of wild-type mouse coilin in knockout cells results in formation of CBs and restores these missing epitopes. Our data demonstrate that full-length coilin is essential for proper formation and/or maintenance of CBs and that recruitment of snRNP and SMN complex proteins to these nuclear subdomains requires sequences within the coilin COOH terminus.