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A unique property of many adult stem cells is their ability to exist in a non-cycling, quiescent state1. Although quiescence serves an essential role in preserving stem cell function until the stem cell is needed in tissue homeostasis or repair, defects in quiescence can lead to an impairment in tissue function2, the extent to which stem cells can regulate quiescence is unknown. Here, we show that the stem cell quiescent state is composed of two distinct functional phases: G0 and an “alert” phase we term GAlert, and that stem cells actively and reversibly transition between these phases in response to injury-induced, systemic signals. Using genetic models specific to muscle stem cells (or satellite cells (SCs)), we show that mTORC1 activity is necessary and sufficient for the transition of SCs from G0 into GAlert and that signaling through the HGF receptor, cMet is also necessary. We also identify G0-to-GAlert transitions in several populations of quiescent stem cells. Quiescent stem cells that transition into GAlert possess enhanced tissue regenerative function. We propose that the transition of quiescent stem cells into GAlert functions as an ‘alerting’ mechanism, an adaptive response that positions stem cells to respond rapidly under conditions of injury and stress without requiring cell cycle entry or a cell fate commitment.

The sonic hedgehog (SHH) signaling pathway directs the embryonic development of diverse organisms and is disrupted in a variety of malignancies. Pathway activation is triggered by binding of hedgehog proteins to the multipass Patched-1 (PTCH) receptor, which in the absence of hedgehog suppresses the activity of the seven-pass membrane protein Smoothened (SMOH). De-repression of SMOH culminates in the activation of one or more of the GLI transcription factors that regulate the transcription of downstream targets. Individuals with germline mutations of the SHH receptor gene PTCH are at high risk of developmental anomalies and of basal-cell carcinomas, medulloblastomas and other cancers (a pattern consistent with nevoid basal-cell carcinoma syndrome, NBCCS). In keeping with the role of PTCH as a tumor-suppressor gene, somatic mutations of this gene occur in sporadic basal-cell carcinomas and medulloblastomas. We report here that a subset of children with medulloblastoma carry germline and somatic mutations in SUFU (encoding the human suppressor of fused) of the SHH pathway, accompanied by loss of heterozygosity of the wildtype allele. Several of these mutations encode truncated proteins that are unable to export the GLI transcription factor from nucleus to cytoplasm, resulting in the activation of SHH signaling. SUFU is a newly identified tumor-suppressor gene that predisposes individuals to medulloblastoma by modulating the SHH signaling pathway through a newly identified mechanism.

(3,4), and two SH2-containing adapter proteins, Grb2 and Shc, have been implicated in its activation. Specifically, these proteins have been shown to bind directly to tyrosine-phosphorylated receptors (5-7) or SH2 docking proteins (such as the insulin receptor substrate 1) (8). Grb2, a 25-kDa protein with two SH3 domains flanking one SH2 domain, shuttles the Ras guanine nucleotide exchange factor, Sosl, to activated receptors (or to insulin receptor substrate 1) (5, 7-11) so that Sosl can activate Ras by catalyzing the exchange of GDP for GTP (5, 9-11). Shc, another widely expressed protein that contains an N-terminal phosphotyrosine binding (PTB) domain (12-16) and a C-terminal SH2 domain (17), can associate, in its tyrosine-phosphorylated form, with Grb2-Sosl complexes and may increase Grb2-Sosl interactions after growth factor stimulation (9,18,19 MATERIALS AND METHODS Reagents. COS-cell-derived murine IL-3 and GM-CSF were produced as described (29). Rabbit antiserum to the Shcassociated p145 protein was generated by immunizing rabbits with a 15-mer obtained from amino acid sequencing (VPAE-GVSSLNEMINP) and crosslinked to keyhole limpet hemocyanin with glutaraldehyde (30). The glutathione S-transferase (GST) fusion proteins, consisting of the 27-kDa N-terminal and of GST linked to the Grb2 N-terminal (amino acid residues 5-56) or C-terminal (residues 163-215) SH3 domains of Grb2, were expressed in Escherichia coli in pGEX-2T plasmids (Pharmacia/LKB) and the fusion proteins were recovered from clarified lysates with glutathione (GSH)-agarose beads as described (31 Abbreviations: GM-CSF, granulocyte-macrophage colony-stimulating factor; GSH, glutathione; GST, glutathione S-transferase; IL-3, interleukin 3; NRS, normal rabbit serum; 5-ptase, inositol polyphosphate 5-phosphatase; Ptd1ns(3,4,5)P3, phosphatidylinositol 3,4,5-trisphosphate; Ins(1,3,4,5)P4, inositol 1,3,4,5-tetrakisphosphate; SH2, src homology 2 domain; PTB, phosphotyrosine binding; p145, the Shc-associated 145-kDa protein.

Summary The ability to maintain quiescence is critical for the long-term maintenance of a functional stem cell pool. To date, the epigenetic and transcriptional characteristics of quiescent stem cells and how they change with age remain largely unknown. In this study, we explore the chromatin features of adult skeletal muscle stem cells, or satellite cells (SCs), which reside predominantly in a quiescent state in fully developed limb muscles of both young and aged mice. Using a ChIP-seq approach to obtain global epigenetic profiles of quiescent SCs (QSCs), we show that QSCs possess a permissive chromatin state in which few genes are epigenetically repressed by Polycomb group (PcG)-mediated histone 3 lysine 27 trimethylation (H3K27me3), and a large number of genes encoding regulators that specify nonmyogenic lineages are demarcated by bivalent domains at their transcription start sites (TSSs). By comparing epigenetic profiles of QSCs from young and old mice, we also provide direct evidence that, with age, epigenetic changes accumulate and may lead to a functional decline in quiescent stem cells. These findings highlight the importance of chromatin mapping in understanding unique features of stem cell identity and stem cell aging.

Satellite cells (SCs) are myogenic stem cells found in skeletal muscle that function to repair tissue damaged by injury or disease. SCs are quiescent at rest, although the signaling pathways required to maintain quiescence are unknown. Using a transgenic Notch reporter mouse and quantitative reverse-transcription polymerase chain reaction analysis of Notch target genes, we determined that Notch signaling is active in quiescent SCs. SC-specific deletion of recombining binding protein-Jκ (RBP-Jκ), a nuclear factor required for Notch signaling, resulted in the depletion of the SC pool and muscles that lacked any ability to regenerate in response to injury. SC depletion was not due to apoptosis. Rather, RBP-Jκ-deficient SCs spontaneously activate, fail to self-renew, and undergo terminal differentiation. Intriguingly, most of the cells differentiate without first dividing. They then fuse with adjacent myofibers, leading to the gradual disappearance of SCs from the muscle. These results demonstrate the requirement of Notch signaling for the maintenance of the quiescent state and for muscle stem cell homeostasis by the regulation of self-renewal and differentiation, processes that are all critical for normal postnatal myogenesis.

This paper describes a personalized k-anonymity model for protecting location privacy against various privacy threats through location information sharing. Our model has two unique features. First, we provide a unified privacy personalization framework to support location k-anonymity for a wide range of users with context-sensitive personalized privacy requirements. This framework enables each mobile node to specify the minimum level of anonymity it desires as well as the maximum temporal and spatial resolutions it is willing to tolerate when requesting for k-anonymity preserving location-based services (LBSs). Second, we devise an efficient message perturbation engine which runs by the location protection broker on a trusted server and performs location anonymization on mobile users' LBS request messages, such as identity removal and spatio-temporal cloaking of location information. We develop a suite of scalable and yet efficient spatio-temporal cloaking algorithms, called CliqueCloak algorithms, to provide high quality personalized location k-anonymity, aiming at avoiding or reducing known location privacy threats before forwarding requests to LBS provider(s). The effectiveness of our CliqueCloak algorithms is studied under various conditions using realistic location data synthetically generated using real road maps and traffic volume data.

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